Silybum marianum cell cultures stably transformed with Vitis vinifera stilbene synthase accumulate t‐resveratrol in the extracellular medium after elicitation with methyl jasmonate or methylated β‐cyclodextrins

The growing demand for t‐resveratrol for industrial uses has generated considerable interest in its production. Heterologous resveratrol production in plant cell suspensions, apart from requiring the introduction of only one or two genes, has the advantage of high biomass yield and a short cultivation time, and thus could be an option for large‐scale production. Silybum marianum is the source of the flavonolignan silymarin. Phenylpropanoid synthesis in cultures of this species can be activated by elicitation with methyl jasmonate and methylated β‐cyclodextrins, with products of the pathway (coniferyl alcohol and some isomers of the silymarin complex) being released into the medium. Given that stilbene synthase shares the same key precursors involved in flavonoid and /or monolignol biosynthesis, we explored the potential of metabolically engineered S. marianum cultures for t‐resveratrol production. Cell suspensions were stably transformed with Vitis vinifera stilbene synthase 3 and the expression of the transgene led to extracellular t‐resveratrol accumulation at the level of milligrams per litre under elicitation. Resveratrol synthesis occurred at the expense of coniferyl alcohol. Production of silymarin was less affected in the transgenic cultures, since the flavonoid pathway is limiting for its synthesis, due to the preferred supply of precursors for the monolignol branch. The fact that the expressed STS gene took excessively produced precursors of non‐bioactive compounds (coniferyl alcohol), while keeping the metabolic flow for target secondary compounds (i.e. silymarin) unaltered, opens a way to extend the applications of plant cell cultures for the simultaneous production of both constitutive and foreign valuable metabolites.     

Ethnopharmacology,Biological Activity and Phytochemistry of Scaevola spinescens

Scaevola spinescens is endemic to Australia and has traditionally been used by Aboriginal and Torres Strait Islander communities to treat a variety of conditions including colds, flu, fever, stomach pain, urinary disorders, sores, tinea, leprosy, and cancer. Extracts prepared from S. spinescens are non-toxic and have been linked with various medicinal properties including antiviral, antibacterial, anti-inflammatory, and anticancer activities. These studies support the ethnopharmacological use of S. spinescens by Indigenous peoples of Australia and highlight the need for further investigations on the plant for potential use in pharmaceutical and food applications. This review provides a comprehensive, up-to-date review of the literature on S. spinescens focusing on the traditional use, medicinal properties, phytochemicals, and factors that affect their composition during pre-treatment and extraction, as well as providing a framework for future studies of the plant.     

Medicinal biotechnology in the genus <Emphasis Type="Italic">scutellaria</Emphasis>

Plant-based medicines have an important role in the lives of millions of people. The ancient knowledge of the use of plants as medicines has led to the discovery of many important western pharmaceuticals, and the popularity of whole plant preparations for a range of therapeutic applications is growing rapidly. However, there are many challenges in the production of plant-based medicines, many of which put both the consumer and the plant populations at risk. Modern biotechnology can be optimized to mass-produce plants of specific chemical composition for use as particular treatments and applications. In this review, we have used one of the most important medicinal plant genera, Scutellaria, as a model to assess the potential of applications of biotechnology for the improvement of medicinal plants.     

Study of different Trichoderma strains on growth characteristics and silymarin accumulation of milk thistle plant

Abstract

Biological control involves the use of beneficial organisms such as Trichoderma that promote positive responses by the plant. Trichoderma species produce and/or release a variety of compounds, including cell wall degrading enzymes and secondary metabolites, which enhance root development, crop productivity and resistance to biotic and abiotic stresses. This study examines the effects of the five Trichoderma strains (M7, KHB, G124-1, G46-3 and G46-7) on milk thistle including morphological characteristics and silymarin accumulation. This research indicated that treating milk thistle by M7 strain, significantly increased the highest branch length of plants, while strain KHB induced the production of silychristin, isosilybin and silymarin. Finally, strain G46-7 dramatically increased silydianin content in milk thistle plant. The findings suggested that certain Trichoderma strains are positive elicitors for promoting growth characteristics and silymarin accumulation in Silybum marianum (L.) Gaertn.     

Are polyamines directly involved in silymarin production in the milk thistle [<Emphasis Type="Italic">Silybum marianum</Emphasis> (L.) Gaernt]?

The possible role of polyamines (PAs) in the regulation of silymarin (Sm) production in milk thistle [Silybum marianum (L.) Gaernt] cell suspension cultures was studied in a young cell culture line (H2 line) and in a synchronized cell line (>3 years; H1 line). The effect of two exogenous PAs, putrescine (Put) and spermidine (Spd), and a number of metabolic inhibitors (L-canavanine, DL-α-difluoromethylornithine, methylglyoxal-bis-guanylhydrazone, cyclohexylamine) on the production of Sm during the growth cycle were analyzed. The results suggest that PAs are not directly involved in the Sm synthesis pathway. In our cell culture system, Sm production and PA contents were determined by the age of the suspension culture cells: with increasing age, the suspension culture cells showed a decreasing capacity to reach the stationary phase during prolonged subculture that was associated with a decreased production of Sm, a steady increase in PA content, and a constant Put/Spd ratio. The synchronization of dividing cells from the S. marianum H1 line did not modify this behaviour. In young cell suspensions, maximum Sm production occurred in the stationary phase, concurrent with the cellular PA contents reaching their minimum value. At the start of the stationary phase, the high percentage of cells in the growth phases (G₀/G₁) and a transient increase in the Put/Spd ratio were accompanied by maximum Sm production and a blockade of cell division.     

Enhanced production of flavonolignans in hairy root cultures of Silybum marianum by over-expression of chalcone synthase gene

In the present study, metabolic engineering approach was used through over-expressing the Petunia chalcone synthase (chsA) gene in order to enhance the silymarin production level in the hairy root cultures of Silybum marianum. Molecular analysis confirmed the presence and integration of chsA transgene in transgenic hairy roots. Chemical analysis indicated that the over-expression of chsA gene enhanced the silymarin production level in the transgenic line as much as 7-folds than the non-transgenic hairy roots. Moreover, the silybin content, the main active component of silymarin, was proved to be 10 times higher in transgenic hairy roots than those of the non-transgenic ones. Therefore, the over-expression of petunia chsA gene in S. marianum hairy roots did not result in gene silencing, but led to an enhanced biosynthesis of the flavonolignans.     

Diversity in plants and other Collembola ameliorate impacts of Sminthurus viridis on plant community structure

Five experiments investigated the importance of herbivory by Sminthurus viridis in structuring botanical composition in developing grasslands, and how these effects may be modified by diversity in collembolan and plant species. Differential susceptibility to S. viridis feeding was demonstrated in 23 dicotyledonous and three monocotyledonous plants assayed as seedlings at the first true leaf stage. The composition of seedling communities developing from natural and artificially constructed soil seed banks varied with the level of S. viridis infestation, with plant species least susceptible to herbivory making the greatest contribution to plant biomass. The combined effect of herbivory by S. viridis and Bourletiella hortensis on Trifolium repens biomass was shown to be less than the effect of S. viridis alone, indicating competitive interference. The adverse effects of herbivory by S. viridis on T. repens biomass was reduced by increased diversity of plants growing in association with the legume, and the presence of four non-herbivorous arthropleonan Collembola. S. viridis was shown to reduce seedling numbers, species diversity and biomass in communities developing from the soil seed bank, but the presence of non-herbivorous arthropleonan species reduced the effect of S. viridis. The experiments demonstrate the potential for herbivory by S. viridis to significantly alter species composition in developing grassland communities. However, interactions with collembolan and plant species profoundly modified S. viridis herbivory impacts, either by reducing feeding intensity or enhancing plant growth. These results highlight the fact that data from simple, synthetic systems may be poor predictors of herbivory impacts under field conditions where more complex species interactions occur.     

Free radical scavenging reactions and antioxidant activities of silybin: Mechanistic aspects and pulse radiolytic studies

Silybin (extracted from Silybum marianum) is the major active constituent of silymarin which possesses a wide range of medicinal properties. These properties may be, in part, due to the potent scavenging capacity of oxidizing free radicals. In this context, scavenging radicals (hydroxyl, azide, dibromide anion radicals, nitrite, carbonate, etc.) of silybin have been studied to understand the mechanistic aspects of its action against free radicals. The transients produced in these reactions have been assigned and the rate constants have been measured by pulse radiolysis techniques. Reduction potential determined both by cyclic voltammetry gave a value 0.62±0.02 V vs NHE at pH 9. Quantum chemical calculations have been performed to further confirm the different activities of individual hydroxyl groups with the difference of heat of formation. Moreover, silybin also protected plasmid pUC18 DNA from soft X-ray radiation which induced strand breaks. These results are expected to be helpful for a better understanding of the anti-oxidative properties of silybin.     

Potential antibacterial and antioxidant inhibitory activities of Silybum marianum mediated biosynthesised He-Ne laser

A potentially beneficial method in laser irradiation is currently gaining popularity. The biosynthesis of low-power lasers has also been applied to the therapy of disease in biological tissues. This study used laser pre-treatments of Silybum marianum (S. marianum) fruit extract as a stabilising agent to bio-fabricate a low-power laser. The silybin A and silybin B of the S. marianum fruit, which are derived from seedlings before S. marianum undergoes therapy with an He-Ne laser at various intervals, were assessed for their expressive properties in this study. The findings revealed that 6-min laser pre-treatments increased silybin A + B and bacterial inhibition and improved the medicinal property of S. marianum. The analysis of the reaction records was performed using ultraviolet–visible spectroscopy. The minimum inhibitory concentration (MIC) limit for the sphere dispersion approach’s antimicrobial effect on the microorganisms under investigation was 50 to 100 g/mL. With an IC50 of 0.69 mg/mL, the laser-treated S. marianum (6 min) demonstrated radical scavenging activity. At MIC concentration, the laser-treated S. marianum (6 min) did not exhibit cytotoxicity in the MCF-7 cell line. Additionally, Salmonella typhi, methicillin-resistant Staphylococcus aureus (MRSA), and E. coli were more susceptible to the antimicrobial effects of ethanolic fruit extract with a greater silybin level. It was observed that the laser-treated S. marianum (6 min) showed beneficial antioxidant and antibacterial properties and could be employed without risk in several medical applications.     

The invasive alien plant species Solidago gigantea alters ecosystem properties across habitats with differing fertility

Question: Invasive alien plants can affect biomass production and rates of biogeochemical cycling. Do the direction and intensity of such effects depend upon the functional traits of native and alien species and upon the properties of the invaded habitat, with the same alien species having differing impacts in different habitats? Location: Lowlands of Switzerland. Methods: Fourteen grassland and wetland sites invaded by Solidago gigantea and widely differing in biomass production and soil P availability were surveyed. To determine whether the impact of the species was related to site fertility, we compared the invaded and native vegetation in terms of biomass, species composition, plant traits and soil properties. Results: S. gigantea generally increased the above‐ground biomass production of the vegetation and soil C content, while reducing nutrient concentrations in biomass and N availability in the soil. However, it had no significant effect on plant species richness, soil respiration, soil pH and P availability. Leaves of S. gigantea had a greater C content than those of native species; other leaf traits and root phosphatase activity did not differ significantly. Conclusions: Our results suggest that a conservative nutrient‐use strategy allows S. gigantea to invade a broad range of habitats. The observed effects of invasion did not vary according to biomass production of the invaded sites, but some effects did depend on soil P availability, being more pronounced at more P‐rich sites. Thus, the full range of invaded habitats should be considered in studying the potential impact of plant invasions on ecosystem processes.     

Assessing the cultivation potential of the energy crop Miscanthus × giganteus for Germany

The European Union in general and Germany in particular want to lead the way to substantially expand renewable energies for power production. Considering the extremely ambitious objectives of the German Federal Government, a strong, nationwide increase in cultivating energy crops can be anticipated. However, the expansion of biomass production, which is already in progress, has led to several environmental and ecological objections. Aside from competing for land, for food and feed production, the expansion of monocultures for biomass and biofuel production with a concentration on maize (Zea mays) and rapeseed (Brassica napus) can be problematic for biodiversity conservation. To face these challenges, the provision and cultivation of additional crop species and cultivars for biomass production would help to avoid these problems. The designated energy crop Miscanthus × giganteus represents an alternative species for extended biomass production. This giant grass is characterized by a broad range of possible applications and a high potential in producing and providing biomass in a sustainable way. In our study, we conducted a Geographic Information System (GIS)-based analysis of the cultivation potential of M. giganteus in Germany. As a result, we generated digital maps that display preferential regions for the cultivation of M. giganteus where a high productivity and quality of biomass is expected. Combining different climate- and soil-dependent scenarios, a total acreage potential of 4 million ha is predicted for Germany.     

Selected Species of the Genus Phellinus – Chemical Composition,Biological Activity,and Medicinal Applications

This study presents the current knowledge on chemical composition, biological activity, and possible medicinal applications of Phellinus igniarius, Phellinus pini, Phellinus pomaceus, and Phellinus robustus. These inedible arboreal species are phytopathogens that cause the enzymatic decomposition of wood. These species belong to the medicinal mushrooms and have been known for centuries in the traditional medicine of the Far East. They have been used as an effective remedy for stomach and intestinal ailments, diarrhea, and hemorrhages. Mycochemical studies have proved the presence of polysaccharides, phenolic compounds, and terpenoids. These compounds show biological activities such as anticancer, antioxidant, antiangiogenic, and antiviral. Research studies conducted using modern analytical methods have advanced the knowledge on the potential therapeutic use of compounds isolated not only from the fruiting bodies but also from biomass obtained with in vitro biotechnological methods.     

Astin C Production by the Endophytic Fungus Cyanodermella asteris in Planktonic and Immobilized Culture Conditions

The fungal endophyte Cyanodermella asteris (C. asteris) has been recently isolated from the medicinal plant Aster tataricus (A. tataricus). This fungus produces astin C, a cyclic pentapeptide with anticancer and anti‐inflammatory properties. The production of this secondary metabolite is compared in immobilized and planktonic conditions. For immobilized cultures, a stainless steel packing immersed in the culture broth is used as a support. In these conditions, the fungus exclusively grows on the packing, which provides a considerable advantage for astin C recovery and purification. C. asteris metabolism is different according to the culture conditions in terms of substrate consumption rate, cell growth, and astin C production. Immobilized‐cell cultures yield a 30% increase of astin C production, associated with a 39% increase in biomass. The inoculum type as spores rather than hyphae, and a pre‐inoculation washing procedure with sodium hydroxide, turns out to be beneficial both for astin C production and fungus development onto the support. Finally, the influence of culture parameters such as pH and medium composition on astin C production is evaluated. With optimized culture conditions, astin C yield is further improved reaching a five times higher final specific yield compared to the value reported with astin C extraction from A. tataricus (0.89 mg g^?1 and 0.16 mg g^?1 respectively).     

Effect of carbohydrates on the production of thaxtomin A by <Emphasis Type="Italic">Streptomyces acidiscabies</Emphasis>

Several Streptomyces species cause plant diseases, including S. scabies, S. acidiscabies and S. turgidiscabies, which produce common scab of potato and similar diseases of root crops. These species produce thaxtomins, dipeptide phytotoxins that are responsible for disease symptoms. Thaxtomins are produced in vivo on diseased potato tissue and in vitro in oat-based culture media, but the regulation of thaxtomin biosynthesis is not understood. S. acidiscabies was grown in a variety of media to assess the impact of medium components on thaxtomin A (ThxA) production. ThxA biosynthesis was not correlated with bacterial biomass, nor was it stimulated by α-solanine or α-chaconine, the two most prevalent potato glycoalkaloids. ThxA production was stimulated by oat bran broth, even after exhaustive extraction, suggesting that specific carbohydrates may influence ThxA biosynthesis. Oat bran contains high levels of xylans and glucans, and both of these carbohydrates, as well as xylans from wheat and tamarind, stimulated ThxA production, but not to the same extent as oat bran. Starches and simple sugars did not induce ThxA production. The data indicate that complex carbohydrates may act as environmental signals to plant pathogenic Streptomyces, allowing production of thaxtomin and enabling bacteria to colonize its host.     

Transport of flavonolignans to the culture medium of elicited cell suspensions of Silybum marianum

Cell suspension cultures of Silybum marianum are able to excrete silymarin compounds into the medium upon elicitation with methyl jasmonate or cyclodextrins. Knowledge of transport mechanism is important to understand Sm metabolism and to develop strategies aimed at increasing production by means of cell cultures. For these reasons, a pharmacological approach was undertaken in this work in order to elucidate the possible mechanism involved in the release of this class of secondary metabolites into the extracellular medium of suspensions.     

Ecophysiological and phytochemical responses of <Emphasis Type="Italic">Salvia sinaloensis</Emphasis> Fern. to drought stress

Salvia sinaloensis Fern. (sage) is a medicinal plant containing plant secondary metabolites (PSMs) with antioxidant properties. The current study investigated the effects of drought stress on S. sinaloensis morphological and ecophysiological traits, and active constituent production. Sage plants were cultivated in controlled conditions for 34 days and exposed to full irrigation as control, half irrigation, or no irrigation. Changes in growth index (G.I.), dry biomass, leaf water potential (LWP), physiological parameters, active compounds, volatilome (BVOCs) and essential oils (EOs) were determined. Not irrigated plants showed a decrease in total chlorophyll content (~???14.7%) and growth (G.I., ~???59.4%) from day 18, and dry biomass at day 21 (??56%), when the complete leaf withering occurred (LWP, ??1.10 MPa). Moderate drought stressed plants showed similar trends for chlorophyll content and growth but kept a constant LWP (??0.35 MPa) and dry biomass throughout the experiment, as control plants. Carotenoids were not affected by water regimes. The photosynthetic apparatus tolerated mild to severe water deficits, without a complete stomatal closure. Plants under both stress conditions increased the percentage of phenols and flavonoids and showed altered BVOC and EO chemical profiles. Interestingly Camphor, the main EO oxygenated monoterpene, increased in moderate stressed plants while the sesquiterpene hydrocarbon Germacrene D decreased. The same trend was seen in the headspace under stress severity. The data evidenced a possible role of the active molecules in the response of S. sinaloensis plants to drought stress. Taking together, these findings point at S. sinaloensis as a potential drought adaptive species, which could be used in breeding strategies to obtain sages with high quality PSMs, saving irrigation water.     

Climate change effects on plant biomass alter dominance patterns and community evenness in an experimental old‐field ecosystem

Atmospheric and climatic change can alter plant biomass production and plant community composition. However, we know little about how climate change‐induced alterations in biomass production affect plant species composition. To better understand how climate change will alter both individual plant species and community biomass, we manipulated atmospheric [CO₂], air temperature, and precipitation in a constructed old‐field ecosystem. Specifically, we compared the responses of dominant and subdominant species to our climatic treatments, and explored how changes in plant dominance patterns alter community evenness over 2 years. Our study resulted in four major findings: (1) all treatments, elevated [CO₂], warming, and increased precipitation increased plant community biomass and the effects were additive rather than interactive, (2) plant species differed in their response to the treatments, resulting in shifts in the proportional biomass of individual species, which altered the plant community composition; however, the plant community response was largely driven by the positive precipitation response of Lespedeza, the most dominant species in the community, (3) precipitation explained most of the variation in plant community composition among treatments, and (4) changes in precipitation caused a shift in the dominant species proportional biomass that resulted in lower community evenness in the wet relative to dry treatments. Interestingly, compositional and evenness responses of the subdominant community to the treatments did not always follow the responses of the whole plant community. Our data suggest that changes in plant dominance patterns and community evenness are an important part of community responses to climatic change, and generally, that such compositional shifts can alter ecosystem biomass production and nutrient inputs.     

Silymarin: an insight to its formulation and analytical prospects

Silymarin, a potential phytochemical compound obtained from the seeds of Silybum marianum plant has been used as a hepatoprotective agent for more than a decade. So far, eight active components of silymarin flavonolignans have been identified, among which silibinin has been proven the most active. However, it had poor oral bioavailability due to extensive phase II metabolism, low permeability across intestinal epithelial cells, low aqueous solubility, and rapid excretion in bile and urine. Therefore it becomes necessary to understand all its formulation and analytical aspects from past to present, including all of its possible future prospects. In modern research scenario, nanotization strategies of drugs has served as a potential approach to enhance solubility, bioavailability and to develop a robust formulation. Several approaches have been utilized previously to enhance the solubility and bioavailability of silymarin to provide it a robust strength against physical, chemical, and environmental degradation. Nanoscale formulations such as nanoemulsion, nanosuspension, liposomes, and solid–lipid nanoparticles can be used to enhance solubility and to target them to desired cells with minimum harm to normal cells. However, many other approaches exist such as dendrimers, ceramic nanoparticles, and carbon nanotubes, which serve as a great vehicle in drug delivery to transport medicament at target sites. Therefore, the purpose of this review was to develop a better understanding of the problems associated with silymarin and approaches to overcome the difficulties to develop a better and stable formulation for food and pharmaceutical applications.     

Effects of water level and temperature on performance of four <Emphasis Type="Italic">Sphagnum</Emphasis> mosses

To evaluate the effects of changes in water level and temperatures on performance of four Sphagnum mosses, S. magellanicum, S. rubellum, S. imbricatum and S. fuscum were grown at two water levels, −5 cm and −15 cm, and at two temperatures, 15°C and 20°C. These species differ in their position along the microtopographical gradient and in their geographical distribution. Height increment, subcapitulum bulk density, biomass production, capitulum water content and cumulative evaporation were measured. Height increment and biomass production of S. magellanicum was lower at low water table than at high water table, whereas height increment and biomass production of S. rubellum, S. imbricatum and S. fuscum were unaffected. Height increment of S. magellanicum, S. rubellum and S. imbricatum was higher at high temperature than at low temperature. Biomass production of only S. magellanicum and S. rubellum was higher at high temperature than at low temperature, corresponding with their more southern distribution. Cumulative evaporation of S. magellanicum and S. rubellum was lower at low water table and could be explained by hampered water transport towards the capitula. We conclude that changes in water table and temperature may alter the Sphagnum composition on raised bogs, which may result in changes to important ecosystem processes. Therefore, it is important that species composition and changes therein are taken into account when evaluating global change effects on raised bog ecosystems.