Clover Root Exudates Favor Novosphingobium sp. HR1a Establishment in the Rhizosphere and Promote Phenanthrene Rhizoremediation.

Rhizoremediation is based on the ability of microorganisms to metabolize nutrients from plant root exudates and, thereby, to cometabolize or even mineralize toxic environmental contaminants. Novosphingobium sp. HR1a is a bacterial strain able to degrade a wide variety of polycyclic aromatic hydrocarbons (PAHs). Here, we have demonstrated that the number of CFU in microcosms vegetated with clover was almost 2 orders of magnitude higher than that in nonvegetated microcosms or microcosms vegetated with rye-grass or grass. Strain HR1a was able to eliminate 92% of the phenanthrene in the microcosms with clover after 9 days. We have studied the molecular basis of the interaction between strain HR1a and clover by phenomic, metabolomic, and transcriptomic analyses. By measuring the relative concentrations of several metabolites exudated by clover both in the presence and in the absence of the bacteria, we identified some compounds that were probably consumed in the rhizosphere; the transcriptomic analyses confirmed the expression of genes involved in the catabolism of these compounds. By using a transcriptional fusion of the green fluorescent protein (GFP) to the promoter of the gene encoding the dioxygenase involved in the degradation of PAHs, we have demonstrated that this gene is induced at higher levels in clover microcosms than in nonvegetated microcosms. Therefore, the positive interaction between clover and Novosphingobium sp. HR1a during rhizoremediation is a result of the bacterial utilization of different carbon and nitrogen sources released during seedling development and the capacity of clover exudates to induce the PAH degradation pathway. IMPORTANCE The success of an eco-friendly and cost-effective strategy for soil decontamination is conditioned by the understanding of the ecology of plant-microorganism interactions. Although many studies have been published about the bacterial metabolic capacities in the rhizosphere and about rhizoremediation of contaminants, there are fewer studies dealing with the integration of bacterial metabolic capacities in the rhizosphere during PAH bioremediation, and some aspects still remain controversial. Some authors have postulated that the presence of easily metabolizable carbon sources in root exudates might repress the expression of genes required for contaminant degradation, while others found that specific rhizosphere compounds can induce such genes. Novosphingobium sp. HR1a, which is our model organism, has two characteristics desirable in bacteria for use in remediation: its ubiquity and the capacity to degrade a wide variety of contaminants. We have demonstrated that this bacterium consumes several rhizospheric compounds without repression of the genes required for the mineralization of PAHs. In fact, some compounds even induced their expression.

Optimization of Two Eco-Friendly Extractions of Black Medick (Medicago lupulina L.) Phenols and Their Antioxidant, Cosmeceutical, α-Glucosidase and α-Amylase Inhibitory Properties.

Medicago lupulina is an ancient edible plant from the Fabaceae family. In this work, two eco-friendly methods for extraction of bioactive phenolics from M. lupulina were developed using mixtures of water with two non-toxic, skin- and environmentally-friendly polyol solvents: glycerol and polypropylene glycol. Ultrasound-assisted extractions were optimized using a Box-Behnken design. The independent variables were the concentration of organic solvent in water (X1), extraction temperature (X2) and time (X3), while the response was phenolic content. The optimum conditions for extraction of polyphenols were (X1, X2, X3): (45%, 70 °C, 60 min) and (10%, 80 °C, 60 min) for glycerol and polypropylene glycol extraction, respectively. The extracts prepared at optimum conditions were rich in phenolic compounds, mainly derivatives of apigenin, kaempferol, luteolin, quercetin, caffeic and ferulic acid, as well as coumestrol. Their cosmeceutical and antidiabetic activity was tested. Both extracts demonstrated notable antioxidant, anti-lipoxygenase and anti-α-amylase activity. In addition to those activities, the glycerol extract efficiently inhibited protein coagulation, elastase and α-glucosidase activity. Glycerol present in the extract displayed enzyme-inhibiting activity in several assays and supported the action of the bioactive constituents. Thus, the optimized glycerol extract is a desirable candidate for direct incorporation in antidiabetic food supplements and cosmeceutical products.

Reduced Water Negatively Impacts Social Bee Survival and Productivity Via Shifts in Floral Nutrition.

Pollinators provide a key ecosystem service vital for the survival and stability of the biosphere. Identifying factors influencing the plant-pollinator mutualism and pollinator management is necessary for maintaining a healthy ecosystem. Since healthy beehives require substantial amounts of carbohydrates (nectar) and protein (pollen) from forage plants such as clover, we must assess how resources offered by plants change under limited water conditions in order to fully understand how drought modifies the pollination mutualism. Here we document how reduced water availability leads to decreased nectar quality and quantity and decreased protein quality of pollen. Furthermore, we provide conclusive evidence that these lower quality resources lead to decreased survival and productivity in both developing honey bees (Hymenoptera: Apidae) and bumble bees (Hymenoptera: Apidae). The results emphasize the importance of the nutritional effects of reduced water on bees when predicting shifts of pollination mutualisms under climate change.

Influence of climate variation on phenolic composition and antioxidant capacity of Medicago minima populations.

Medicago minima is a pasture legume that grows almost all over the world. In Tunisia, it occupies various climatic environments and is considered the most abundant annual Medicago plant. However, this species is unconsumed and unused by humans. This study aimed to explore the phytochemical characteristics of Medicago minima selected from different provenances in Tunisia and subsequently investigate the influence of environmental factors on their phenolic composition and antioxidant activity. Therefore, a calorimetric method and DPPH tests provided the total phenolic and total flavonoid contents and antioxidant potential in roots, stems, leaves and seeds. High performance liquid chromatography (HPLC) identified and quantified four phenolic acids and three flavonoids in the studied organs. Roots and leaves showed the greatest phenolic compound content and had high antioxidant activity. Rutin and syringic acid (leaves) represent a characteristic for this species. For each organ, principal component analysis of phenolic profiles showed that the root's phenolic composition could be an indication of the plant adaptation to even small changes in its environments. Plants originating from a cold climate, higher altitude or semi-arid environment had the highest phenolic compound contents in their organs. Our findings provide useful information for the exploitation of the phenolic compounds in these weeds for the development of environmental sustainability.

Membrane Extracts from Plant Tissues.

The comparison of isolated plant cell membranous enclosures can be hampered if their extraction method differs, e.g., in regard to the utilized buffers, the tissue, or the developmental stage of the plant. Thus, for comparable results, different cellular compartments should be isolated synchronously in one procedure. Here, we devise a workflow to isolate different organelles from one tissue, which is applicable to different eudicots such as Medicago x varia and Solanum lycopersicum. We describe this method for the isolation of different organelles from one plant tissue for the example of Arabidopsis thaliana. All compartments are retrieved by utilizing differential centrifugation with organelle-specific parameters.

In Vitro Anthelmintic Activity of Saponins from Medicago spp. Against Sheep Gastrointestinal Nematodes.

Gastrointestinal strongyle nematodes (GIS) are included among the most important parasites of small ruminants. The widespread drug resistance and drug residues in products of animal origin have increased the interest in the search for natural compounds with anthelmintic activity as a valid alternative to current synthetic drugs. The aim of the present investigation was to test the 'in vitro' anthelmintic activity of saponins and prosapogenins from different Medicago species, selected for their importance as a forage crop worldwide for animal feeding. From these plants, saponin mixtures were extracted, purified and used at scalar concentrations to evaluate their anthelmintic activities against sheep gastrointestinal strongyles (GISs), by the egg hatch test. Treated and untreated controls were used as the comparison. Data were statistically analyzed, and EC50 and EC90 were also calculated. All saponins and prosapogenins showed inhibiting effects on GIS eggs in a concentration-dependent manner. At higher concentrations, most of them showed an efficacy comparable to the reference drug (Thiabendazole 3 µg/mL) (P < 0.001). With 1.72 mg/mL EC50 and 3.84 mg/mL EC90, saponin from M. polymorpha cultivars Anglona was the most active. Obtained results encourage further studies aimed at evaluating the efficacy 'in vivo' of saponins which resulted as most effective 'in vitro' in this study.

Phytochemical and cytotoxic evaluation of Medicago monantha: In vivo protective potential in rats.

This research focuses on screening and evaluation of bioactive constituents in plants through pharmacological assays. In present study, we evaluated phytochemicals, cytotoxic activity, in vivo effect of M. monantha against CCl4 induced toxicity in cardiac and renal tissues and its aphrodisiac potential in rats. Shade dried plant was extracted with methanol. The phytochemical screening indicates the presence of flavonoids and alkaloids. Aphrodisiac study showed improved sexual desire; may be attributed to the presence of saponins that boosts the androgen level. Cytotoxicity of the plant was assessed through brine shrimp lethality assay and nearly all the fractions showed promising results. The in vivo study focused on the protective ability of extract against CCl4-induced oxidative damage in renal and cardiac tissues of rats. Serum analysis revealed that CCl4 intoxication increased the levels of bilirubin and blood urea nitrogen (BUN). Antioxidant enzyme analysis showed that catalase, peroxidase, superoxide dismutase, glutathione-S-transferase, glutathione activity and protein levels declined due to CCl4 induced renal and cardiac toxicity. Moreover, the histopathological studies of both low & high dose plant treated group's revealed glomerular hypertrophy and glomerular congestion in kidney, cardiac degeneration and vacuolization of germinal epithelium induced by CCl4 intoxication. DNA also shows damage showed the toxic nature of the plant.

Effects of Magnesium Fertilizer on the Forage Crude Protein Content Depend upon Available Soil Nitrogen.

Magnesium (Mg) is important for both plant photosynthesis and protein synthesis. Nevertheless, latent Mg deficiencies are common, and Mg addition has shown an improved yield. Might such an increasing yield cause "hidden" hunger for microelements and protein, and if so, what is the mechanism? We conducted two greenhouse experiments using low-Mg soil to investigate (i) effects of five levels of Mg fertilizer (20-400 mg kg-1) on eight elements and crude protein concentrations in annual ryegrass and white clover and (ii) if any protein effects of the Mg fertilizer depend upon soil nitrogen (N). Mg addition significantly increased the yield in both species, simultaneously decreasing concentrations of crude protein, calcium (Ca), sodium, manganese, and potassium/Mg and Ca/Mg ratios caused by increased biomass dilution effects and increased [Mg]. Other mineral dilution effects of the Mg fertilizer depended upon species: the concentration of phosphorus decreased only in ryegrass, and the concentration of zinc decreased only in white clover. Mg addition in soil rich with available N (from N fertilizer in ryegrass or biological fixation in white clover) showed an increased crude protein content as well as increased yield in the forage of both species. These results suggest that the Mg fertilizer can affect the protein content positively or negatively depending upon available N in soil and that sufficiently available N must be ensured along with Mg addition in low Mg soils.

A laser optical method for predicting adulteration of clover honey.

BACKGROUND: The authenticity of honey is of great importance for commercial and health reasons, because adulteration of bee honey with cheaper sweetening materials is sometimes carried out for financial gain. The objectives of this study were to evaluate clover honey quality according to physicochemical characteristics, to measure light transmission through honey samples using a visible laser technique (He-Ne laser at 632.8 nm) and to predict types of adulterants of clover honey according to the transmission intensity of the laser beam through the sample. RESULTS: The transmission intensity of pure clover honey was 3520 lx. Addition of water or sugar solution at various concentrations led to increased transmission intensities in the range 4720-6980 lx, while addition of glucose honey led to increased transmission intensities in the range 3570-4120 lx. However, addition of starch suspension or black honey led to decreased transmission intensities in the ranges 3440-120 and 2770-85 lx respectively. The physicochemical properties of pure clover honey were within the standard limits. The light microscope image of pollen in the honey indicated that the pollen came from the clover plant. CONCLUSION: The transmission intensity method using visible laser light to predict the type of adulteration of clover honey can be considered a fast and accurate method. © 2018 Society of Chemical Industry.

Small-Seeded Legumes as a Novel Food Source. Variation of Nutritional, Mineral and Phytochemical Profiles in the Chain: Raw Seeds-Sprouted Seeds-Microgreens.

Growing public concerns about health haves prompted the search for novel food sources. The study is focused on the seeds, sprouted seeds and microgreens of Trifolium pratense, T. medium, Medicago sativa, M. lupulina, Onobrychis viciifolia, Astragalus glycyphyllos and A. cicer species as a potential source of value-added food ingredientsr. The samples were analysed for nutritional (wet chemistry, standard methods) and mineral (atomic absorption spectroscopy, UV-Vis spectrophotometry) profiles, isoflavones (ultra-performance liquid with diode array detector ⁻UPLC-DAD), coumestrol (UPLC-DAD), condensed tannins (CT) (vanillin-H2SO4 assay) and triterpene saponins (UPLC with triple-stage quadrupole MS). In our study, each species displayed high, but species-dependent nutritional, mineral and phytochemical value. All counterparts of legumes were mineral and protein rich. A. glycyphyllos samples, especially seeds, were abundant in iron. Trifolium spp. were found to be important sources of isoflavones, Medicago spp. of coumestrol and saponins, and O. viciifolia of CT. The protein and phytochemical contents increased and total carbohydrates decreased from seeds to microgreens.Our findings proved for the first time that seeds, sprouted seeds, and especially microgreens of small-seeded legumes are promising new sources of ingredients for fortification of staple foods with bioactive compounds, minerals and nutrients.

Input of different kinds of soluble pectin to cation binding properties of roots cell walls.

It is widely believed that pectin are responsible for the vast majority of cation binding positions in the root cell walls. To estimate the role of particular kinds of pectin, we studied the cell wall material isolated from the roots of monocots (wheat and rye) and dicots (clover and lupine) before and after removal of different fractions of soluble pectin. Simultaneously PME activity and degree of pectin methylation were determined. From potentiometric titration curves cation exchange capacity, total surface charge and acidic strength of surface functional groups responsible for surface charging were determined. Monocots had smaller cation exchange capacity and lower pectin content than dicots. Removal of pectin induced up to 50% reduction in the cell walls surface charge. Pectin seem to have more acidic character than the other roots components that is seen from an increase in very weakly acidic groups fraction and significant decrease in the average dissociation constant of the cell walls material after pectin removal. Water soluble pectin and non-pectic soluble compounds had the dominant role in surface charging, while chelator and diluted alkali soluble pectin contributed to surface charge only at high pH's.

Activity of Saponins from Medicago species Against HeLa and MCF-7 Cell Lines and their Capacity to Potentiate Cisplatin Effect.

BACKGROUND: Saponins from Medicago species display several biological activities, among them apoptotic effects against plant cells have been evidenced. In contrast, their cytotoxic and antitumor activity against animal cells have not been studied in great details. OBJECTIVE: To explore the cytotoxic properties of saponin from Medicago species against animal cells and their effect in combination with the antitumoral drug cisplatin. METHOD: Cytotoxic activity of saponin mixtures from M. arabica (tops and roots), M. arborea (tops) and M. sativa (tops, roots and seeds) and related prosapogenins from M. arborea and M. sativa (tops) against HeLa and MCF-7 cell lines is described. In addition, cytotoxicity of soyasaponin I and purified saponins (1-8) of hederagenin, medicagenic and zanhic acid is also presented. Combination experiments with cisplatin have been also conducted. RESULTS: Saponins from M. arabica tops and roots (mainly monodesmosides of hederagenin and bayogenin) were the most effective to reduce proliferation of HeLa and MCF-7 cell lines. Among the purified saponins, the most cytotoxic was saponin 1, 3-O-ß-D-glucopyranosyl(1→2)-α-L-arabinopyranosyl hederagenin. When saponins, derived prosapogenins and pure saponins were used in combination with cisplatin, they all, to different extent, were able to potentiate cisplatin activity against HeLa cells but not against MCF-7 cell lines. Moreover uptake of cisplatin in these cell lines was significantly reduced. CONCLUSION: Overall results showed that specific molecular types of saponins (hederagenin glycosides) have potential as anti-cancer agents or as leads for anti-cancer agents. Moreover saponins from Medicago species have evidenced interesting properties to mediate cisplatin effects in tumor cell lines.

Combined Red Clover isoflavones and probiotics potently reduce menopausal vasomotor symptoms.

BACKGROUND: Natural estrogen decline leads to vasomotor symptoms (VMS). Hormone therapy alleviates symptoms but increases cancer risk. Effective treatments against VMS with minimal cancer risks are needed. We investigate the effects of a highly bioavailable aglycone rich Red Clover isoflavone treatment to alleviate existing menopausal VMS, assessed for the first time by 24hour ambulatory skin conductance (SC). METHODS AND RESULTS: We conducted a parallel, double blind, randomised control trial of 62 peri-menopausal women aged 40-65, reporting ≥ 5 hot flushes/day and follicle stimulating hormone ≥35 IU/L. Participants received either twice daily treatment with bioavailable RC extract (RCE), providing 34 mg/d isoflavones and probiotics, or masked placebo formulation for 12 weeks. The primary outcome was change in daily hot flush frequency (HFF) from baseline to 12 weeks using 24hr SC. Secondary outcomes were change in SC determined hot flush intensity (HFI), self-reported HFF (rHFF) and hot flush severity (rHFS), blood pressure and plasma lipids. A significant decrease in 24hr HFF (P < 0.01) and HFI (P<0.05) was found when comparing change from baseline to 12 months of the RCE (-4.3 HF/24hr, CI -6.8 to -2.3; -12956 µS s-1, CI -20175 to -5737) with placebo (0.79 HF/24hr, CI -1.56 to 3.15; 515 µS s-1, CI -5465 to 6496). rHFF was also significantly reduced (P <0.05)in the RCE (-2.97 HFs/d, CI -4.77 to -1.17) group compared to placebo (0.036 HFs/d, CI -2.42 to 2.49). Other parameters were non-significant. RCE was well tolerated. CONCLUSION: Results suggest that moderate doses of RCE were more effective and superior to placebo in reducing physiological and self-reported VMS. Findings support that objective physiological symptom assessment methods should be used together with self-report measures in future studies on menopausal VMS. TRIAL REGISTRATION: ClinicalTrials.gov NCT02028702.

Artefact formation during acid hydrolysis of saponins from Medicago spp.

Artefact compounds obtained during acid hydrolysis of saponins from Medicago spp. (Fabaceae), have been monitored and evaluated by GC-FID. Their identification has been performed by GC-MS and 1H and 13C NMR. Saponins with different substituents on the triterpenic pentacyclic aglycones were considered, and their hydrolysis products were detected and quantified during 10 h of time course reaction. From soyasapogenol B glycoside the well known soyasapogenols B, C, D and F were obtained together with a previously undescribed sapogenol artefact identified as 3ß,22ß,24-trihydroxyolean-18(19)-en and named soyasapogenol H. From a zanhic acid saponin two major artefact compounds identified as 2ß,3ß,16α-trihydroxyolean-13(18)-en-23,28-dioic acid and 2ß,3ß,16α-trihydroxyolean-28,13ß-olide-23-oic acid were obtained, together with some zanhic acid. Other compounds, detected in very small amount in the reaction mixture, were also tentatively identified based on their GC-MS and UV spectra. The other most characteristic saponins in Medicago spp., hederagenin, bayogenin and medicagenic acid glycosides, under acidic condition of hydrolysis, released instead the correspondent aglycones and generated a negligible amount of artefacts. Nature of artefacts and mechanism of their formation, involving a stable tertiary carbocation, is here proposed and discussed for the first time.

Whole lactation production responses in high-yielding dairy cows using high-quality grass/clover silage.

BACKGROUND: Limiting the use of purchased concentrate for livestock and replacing it with home-grown forage without compromising milk production can offer benefits in both organic and conventional dairy systems. A full lactation trial was conducted with 92 cows over two years comparing three diets, each differing in the mean forage proportion over the lactation, 500 (500F), 600 (600F) and 700 (700F) g kg-1 dry matter (DM) respectively. The diets were designed to represent common conventional feeding, current regulations for organic production and more extreme high-forage-based production respectively. The aims were to determine the effects of forage proportion in the diet on milk production and feed utilisation. RESULTS: Compared with 500F, daily milk yield did not differ in 600F but was lower in 700F (31.3, 31.1 and 29.2 kg energy-corrected milk respectively). Daily dry matter intake (DMI) was similar between treatments (20.3, 20.4 and 19.9 kg in 500F, 600F and 700F respectively). CONCLUSION: Increasing the forage proportion from 500 to 600 g kg-1 DM did not have any adverse effects on milk production or DMI. Thus it is possible to produce the same quantity of milk with less concentrate and reduce the use of potential human feeds in dairy production. © 2016 Society of Chemical Industry.

Leguminose green juice as an efficient nutrient for l(+)-lactic acid production.

Lactic acid is one of the most important building blocks for the production of bioplastic. Many investigations have been conducted to reduce the lactic acid production costs. In this work, the focus was put on the application of legume pressed juice or green juice as nutrient source. The pressed juice was utilized directly without prior pre-treatment and sterilization. Using two different alfalfa green juices and a clover green juice from two different harvest years as sole nutrients, non-sterile fermentations were performed at 52°C and pH 6.0 with a thermotolerant strain Bacillus coagulans AT107. The results showed that alfalfa green juices generally were more suitable for high lactic acid production than clover green juices, presumably due to the higher nitrogen content. A final titer of 98.8g/L after 30h with l(+)-lactic acid purity of >99% was obtained.

Screening of saponins and sapogenins from Medicago species as potential PPARγ agonists and X-ray structure of the complex PPARγ/caulophyllogenin.

A series of saponins and sapogenins from Medicago species were tested for their ability to bind and activate the nuclear receptor PPARγ by SPR experiments and transactivation assay, respectively. The SPR analysis proved to be a very powerful and fast technique for screening a large number of compounds for their affinity to PPARγ and selecting the better candidates for further studies. Based on the obtained results, the sapogenin caulophyllogenin was proved to be a partial agonist towards PPARγ and the X-ray structure of its complex with PPARγ was also solved, in order to investigate the binding mode in the ligand binding domain of the nuclear receptor. This is the first known crystal structure of a sapogenin directly interacting with PPARγ. Another compound of the series, the echinocistic acid, showed antagonist activity towards PPARγ, a property that could be useful to inhibit the adipocyte differentiation which is a typical adverse effect of PPARγ agonists. This study confirms the interest on saponins and sapogenins as a valuable natural resource exploitable in the medical and food industry for ameliorating the metabolic syndrome.

Historical nectar assessment reveals the fall and rise of floral resources in Britain.

There is considerable concern over declines in insect pollinator communities and potential impacts on the pollination of crops and wildflowers. Among the multiple pressures facing pollinators, decreasing floral resources due to habitat loss and degradation has been suggested as a key contributing factor. However, a lack of quantitative data has hampered testing for historical changes in floral resources. Here we show that overall floral rewards can be estimated at a national scale by combining vegetation surveys and direct nectar measurements. We find evidence for substantial losses in nectar resources in England and Wales between the 1930s and 1970s; however, total nectar provision in Great Britain as a whole had stabilized by 1978, and increased from 1998 to 2007. These findings concur with trends in pollinator diversity, which declined in the mid-twentieth century but stabilized more recently. The diversity of nectar sources declined from 1978 to 1990 and thereafter in some habitats, with four plant species accounting for over 50% of national nectar provision in 2007. Calcareous grassland, broadleaved woodland and neutral grassland are the habitats that produce the greatest amount of nectar per unit area from the most diverse sources, whereas arable land is the poorest with respect to amount of nectar per unit area and diversity of nectar sources. Although agri-environment schemes add resources to arable landscapes, their national contribution is low. Owing to their large area, improved grasslands could add substantially to national nectar provision if they were managed to increase floral resource provision. This national-scale assessment of floral resource provision affords new insights into the links between plant and pollinator declines, and offers considerable opportunities for conservation.

Therapeutic Potential of Temperate Forage Legumes: A Review.

The discovery of bioactive molecules from botanical sources is an expanding field, preferentially oriented to plants having a tradition of use in medicine and providing high yields and availability. Temperate forage legumes are Fabaceae species that include worldwide-important crops. These plants possess therapeutic virtues that have not only been used in veterinary and folk medicine, but have also attracted the interest of official medicine. We have examined here Medicago sativa (alfalfa), Trifolium pratense and T. repens (clovers), Melilotus albus and M. officinalis (sweet clovers), Lotus corniculatus (birdsfoot trefoil), Onobrychis viciifolia (sainfoin), Lespedeza capitata (roundhead lespedeza), and Galega officinalis (goat's rue). The phytochemical complexes of these species contain secondary metabolites whose pharmacological potentials deserve investigation. Major classes of compounds include alkaloids and amines, cyanogenic glycosides, flavonoids, coumarins, condensed tannins, and saponins. Some of these phytochemicals have been related to antihypercholesterolemia, antidiabetic, antimenopause, anti-inflammatory, antiedema, anthelmintic, and kidney protective effects. Two widely prescribed drugs have been developed starting from temperate forage legumes, namely, the antithrombotic warfarin, inspired from sweet clover's coumarin, and the antidiabetic metformin, a derivative of sainfoin's guanidine. Available evidence suggests that temperate forage legumes are a potentially important resource for the extraction of active principles to be used as nutraceuticals and pharmaceuticals.

Biogas potential from forbs and grass-clover mixture with the application of near infrared spectroscopy.

This study investigated the potentials of forbs; caraway, chicory, red clover and ribwort plantain as substrates for biogas production. One-, two- and four-cut systems were implemented and the influence on dry matter yields, chemical compositions and methane yields were examined. The two- and four-cut systems resulted in higher dry matter yields (kg [total solid, TS] ha(-1)) compared to the one-cut system. The effect of plant compositions on biogas potentials was not evident. Cumulative methane yields (LCH4kg(-1) [volatile solid, VS]) were varied from 279 to 321 (chicory), 279 to 323 (caraway), 273 to 296 (ribwort plantain), 263 to 328 (red clover) and 320 to 352 (grass-clover mixture), respectively. Methane yield was modelled by modified Gompertz equation for comparison of methane production rate. Near infrared spectroscopy showed potential as a tool for biogas and chemical composition prediction. The best prediction models were obtained for methane yield at 29 days (99 samples), cellulose, acid detergent fibre, neutral detergent fibre and crude protein, (R(2)>0.9).