Plant components with specific activities against rumen methanogens.

A wide range of plant bioactive components (phytochemicals) have been identified as having potential to modulate the processes of fermentation in the rumen. The use of plants or plant extracts as natural feed additives has become a subject of interest not only among nutritionists but also other scientists. Although a large number of phytochemicals (e.g. saponins, tannins and essential oils) have recently been investigated for their methane reduction potential, there have not yet been major breakthroughs that could be applied in practice. A key tenet of this paper is the need for studies on the influence of plant components on methane production to be performed with standardized samples. Where there are consistent effects, the literature suggests that saponins mitigate methanogenesis mainly by reducing the number of protozoa, condensed tannins both by reducing the number of protozoa and by a direct toxic effect on methanogens, whereas essential oils act mostly by a direct toxic effect on methanogens. However, because the rumen is a complex ecosystem, analysis of the influence of plant components on the populations of methanogens should take into account not only the total population of methanogens but also individual orders or species. Although a number of plants and plant extracts have shown potential in studies in vitro, these effects must be confirmed in vivo.

Dietary plant bioactives for poultry health and productivity.

1. Plants and their biologically active chemical constituents, sometimes called secondary metabolites or bioactives, present numerous opportunities for the improvement of livestock production by inclusion in the diet. 2. Many such plant derived materials have well established therapeutic values in man; however, their potential as feed additives in animal production, particularly of poultry, remains largely unexploited. 3. There is increasing evidence indicating that they can be efficient in controlling diseases, and plant bioactives may also influence production parameters such as feed efficiency and product quality. 4. It has been reported that they may even replicate some of the effects of antibiotic growth promoters, which were banned from use in Europe from 2006. 5. This review assesses the status of plant bioactives in poultry production and their mode of action on avian physiology, particularly in the digestive tract.

Studies on antioxidant properties of polyphenol-rich extract from berries of Aronia melanocarpa in blood platelets.

The antioxidant properties of extract from berries of Aronia melanocarpa (chokeberry) containing: anthocyanidines, phenolic acids and quercetine glycosides on oxidative/nitrative stress induced by peroxynitrite (ONOO(-), a powerful physiological oxidant, nitrating species and inflammatory mediator) in human blood platelets were studied in vitro. The extract from A. melanocarpa (5 - 50 microg/mL) significantly inhibited platelet protein carbonylation (measured by ELISA method) and thiol oxidation estimated with 5,5'-dithio-bis(2-nitro-benzoic acid) (DTNB) induced by peroxynitrite (0.1 mM) (IC(50)--35 microg/mL for protein carbonylation, and IC(50)--33 microg/mL for protein thiol oxidation). The tested extract only slightly reduced platelet protein nitration (measured by C- ELISA method). The extract also caused a distinct reduction of platelet lipid peroxidation induced by peroxynitrite. Moreover, in our preliminary experiments we observed that the extract (50 microg/mL) reduced oxidative/nitrative stress in blood platelets from patients with breast cancer. The obtained results indicate that in vitro the extract from A. melanocarpa has the protective effects against peroxynitrite-induced oxidative/nitrative damage to the human platelet proteins and lipids. The extract from A. melanocarpa seems to be also useful as an antioxidant in patients with breast cancer.

Anti-inflammatory and anti-arthritic effects of Yucca schidigera: a review.

Yucca schidigera is a medicinal plant native to Mexico. According to folk medicine, yucca extracts have anti-arthritic and anti-inflammatory effects. The plant contains several physiologically active phytochemicals. It is a rich source of steroidal saponins, and is used commercially as a saponin source. Saponins have diverse biological effects, including anti-protozoal activity. It has been postulated that saponins may have anti-arthritic properties by suppressing intestinal protozoa which may have a role in joint inflammation. Yucca is also a rich source of polyphenolics, including resveratrol and a number of other stilbenes (yuccaols A, B, C, D and E). These phenolics have anti-inflammatory activity. They are inhibitors of the nuclear transcription factor NFkappaB. NFkB stimulates synthesis of inducible nitric oxide synthase (iNOS), which causes formation of the inflammatory agent nitric oxide. Yucca phenolics are also anti-oxidants and free-radical scavengers, which may aid in suppressing reactive oxygen species that stimulate inflammatory responses. Based on these findings, further studies on the anti-arthritic effects of Yucca schidigera are warranted.

Chromatographic determination of plant saponins--an update (2002-2005).

The developments during 2002-2005 in the methods used for saponin analyses in plant material are presented. There were number of papers published on isolation and identification of new saponins by chromatographic techniques. Some new developments can be found in separation techniques or solid and mobiles phases used. Separation of individual saponins is still complicated and time consuming. This is due to the fact that in most of the plant species saponins occur as a multi-component mixture of compounds of very similar polarities. Thus, to isolate single compound for structure elucidation or biological activity testing, a combination of different chromatographic techniques has to be used, e.g. first separation of the mixture to simpler sub-fractions on reversed phase C18 has to be followed by further purification on normal phase Silica gel column. Especially difficult is determination of saponins in plant material as these compounds do not possess chromophores and their profiles cannot be registered in UV. Most HPLC methods apply not only specific registration at 200-210 nm, but these methods are not applicable for determination of many saponins in plant material at levels lower than 200-300 mg/kg. Some new or improved techniques for quantification of saponins in plant material were published in reviewed period. These include further progress in the application of evaporative light scattering detection (ELSD) for saponin profiling and quantification, which is also not only specific but also more sensitive in comparison to 200-210 nm detection. Some progress in development of new applications for liquid chromatography-electrospray mass spectrometry (LC/ESI/MS) for saponin determination has also been done. This method gives highest sensitivity and on line identification of separated saponins and should be recommended for specialized analyses of extracts and pharmaceutical formulas like the validation of a new assay. From non-chromatographic techniques for saponin determination, a sensitive and compound specific ELISA tests for some saponins were developed.

First European interlaboratory study of the analysis of benzoxazinone derivatives in plants by liquid chromatography.

Six laboratories from four different countries participated in the first European interlaboratory comparison exercise within the framework of the "Fate and toxicity of allelochemicals (natural plant toxins) in relation to environment and consumer" (FATEALLCHEM) European Union Project. The study, organized between November 2002 and March 2003, involved the analyses of seven benzoxazinone derivatives in two standard solutions and one purified extract of root material. Results are reported from the first phase of the study that examined the variability associated with different detection methods and different laboratories. The analytical strategies were based on liquid chromatography (LC) with diode array detection, LC coupled to mass spectrometry (MS) and LC coupled to tandem MS. When data from all laboratories were pooled, the relative standard deviation values ranged from 2 to 14% for the determination of target compounds in standard solutions, and between 19 and 47% for the analysis in root material. Comparison of the three detection techniques leads to the conclusion that MS approaches are the most accurate and precise techniques for the determination of benzoxazinone derivatives at ng/microL level in plant material.

Chromatographic determination of plant saponins.

The methods used for saponin determination in plant materials are presented. It is emphasised that the biological and spectrophotomeric methods still being used for saponin determination provide, to some extent, valuable results on saponin concentrations in plant material. However, since they are sensitive to the structural variation of individual saponins they should be standardized with saponin mixtures isolated from the plant species in which the concentration is measured. However, one plant species may contain some saponins which can be determined with a biological test and others which cannot. That is why biological and colorimetric determinations do not provide accurate data and have to be recognized as approximate. Thin-layer chromatography on normal and reversed-phases (TLC, HPTLC, 2D-TLC) provides excellent qualitative information and in combination with on-line coupling of a computer with dual-wavelength flying-spot scanner and two-dimensional analytical software can be used for routine determination of saponins in plant material. The densitometry of saponins has been very sensitive, however, to plate quality, spraying technique and the heating time and therefore appropriate saponin standards have to be run in parallel with the sample. Gas-liquid chromatography has limited application for determination since saponins are quite big molecules and are not volatile compounds. Thus, there are only few applications of GC for determination of intact saponins. The method has been used for determination of TMS, acetyl or methyl derivatives of an aglycones released during saponin hydrolysis. However, structurally different saponins show different rates of hydrolysis and precise optimisation of hydrolysis conditions is essential. Besides, during hydrolysis a number of artefacts can be formed which can influence the final results. High performance liquid chromatography on reversed-phase columns remains the best technique for saponin determination and is the most-widely used method for this group of compounds. However, the lack of chromophores allowing detection in UV, limits the choice of gradient and detection method. The pre-column derivatisation with benzoyl chloride, coumarin or 4-bromophenacyl bromide has been used successfully in some cases allowing UV detection of separation. Standardisation and identification of the peaks in HPLC chromatograms has been based on comparison of the retention times with those observed for authentic standards. But new hyphenated techniques, combining HPLC with mass spectrometry and nuclear magnetic resonance are developing rapidly and allow on-line identification of separated saponins. Capillary electrophoresis has been applied for saponin determination only in a limited number of cases and this method is still being developed.

Alfalfa (Medicago sativa L.) flavonoids. 2. Tricin and chrysoeriol glycosides from aerial parts.

Ten flavone glycosides have been isolated and identified in aerial parts of alfalfa. These included six tricin, one 3'-O-methyltricetin, and three chrysoeriol glycosides. Most of these compounds were acylated with ferulic, coumaric, or sinapic acids, and acylation occurred on the terminal glucuronic acid. Eight of these compounds, including 7-O-beta-D-glucuronopyranosyl-3'-O-methyltricetin, 7-O-beta-D-glucuronopyranosyl-4'-O-beta-D-glucuronopyranosidechrysoeriol, 7-O-[2'-O-feruloyl-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]chrysoeriol, 7-O-[2'-O-feruloyl-[beta-D-glucuronopyranosyl(1-->3)]-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]chrysoeriol, 7-O-[2'-O-sinapoyl-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]tricin, 7-O-[2'-O- feruloyl-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]tricin, 7-O-[2'-O-p-coumaroyl-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]tricin, and 7-O-[2'-O-feruloyl-[beta-D-glucuronopyranosyl(1-->3)]-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]tricin, have not been reported previously in the plant kingdom. Two previously identified alfalfa flavones, 7-O-beta-D-glucuronopyranosidetricin and 7-O-[beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]tricin, were also isolated.

Steroidal saponins of Yucca schidigera Roezl.

Eight steroidal saponins have been isolated from Yucca schidigera Roezl. trunk, and their structures were established by spectral (MS and NMR) techniques. These included three novel furostanol glycosides including 3-O-beta-D-glucopyranosyl-(1-->2)-[beta-D-xylopyranosyl-(1-->3)]-beta-D-glucopyranosyl-5 beta(25R)-furostan-3 beta,22 alpha,26-triol 26-O-beta-D-glucopyranoside, 3-O-beta-D-glcopyranosyl-(1-->2)-[beta-D-xylopyranosyl-(1-->3)]-beta-D-glucopyranosyl-5 beta(25R)-furost-20(22)-en-3 beta,26-diol-12-one 26-O-beta-D-glucopyranoside, 3-O-beta-D-glcopyranosyl-(1-->2)-beta-D-glucopyranosyl-5 beta(25R)-furostan-3 beta,22 alpha,26-triol 26-O-beta-D-glucopyranoside, and five known spirostanol glycosides. On the basis of the extraction efficiency, furostanol glycosides made up only 6.8% of total saponins isolated.

Acylated apigenin glycosides from alfalfa (Medicago sativa L.) var. Artal.

Three flavones, including 4'-O-[2'-O-E-feruloyl-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]apigenin, 7-O-beta-D-glucuronopyranosyl-4'-O-[2'-O-E-feruloyl-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]apigenin and 7-O-beta-D-glucuronopyranosyl-4'-O-[2'-O-p-E-coumaroyl-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]apigenin have been identified in alfalfa var. Artal. The known flavone 7-O-[2-O-E-feruloyl-[beta-D-glucuronopyranosyl(1-->3)]-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucurono-pyranoside] apigenin was also isolated. The structures of these compounds were deduced on the basis of their spectral data.

Antioxidant and antiradical activities of flavonoids.

The relationship between the structure of 42 flavonoids and their antioxidant and antiradical activities was elucidated by heat-induced oxidation in a beta-carotene and linoleic acid system and by the 1,1-diphenyl-2-picrylhydrazyl decoloration test. From seven structurally divergent groups of flavonoids, only flavonols with a free hydroxyl group at the C-3 position of the flavonoid skeleton showed high inhibitory activity to beta-carotene oxidation. Antiradical activity depended on the presence of a flavonol structure or free hydroxyl group at the C-4' position. The effect of the 4'-hydroxyl was strongly modified by other structural features, such as the presence of free hydroxyls at C-3 and/or C-3' and a C2-C3 double bond.

Resveratrol and other phenolics from the bark of Yucca schidigera roezl.

Five phenolic constituents have been identified in Yucca schidigera bark, and their structures were established by spectral (FABMS and NMR) experiments. These included two known stilbenes, trans-3,4',5-trihydroxystilbene (resveratrol) and trans-3,3',5,5'-tetrahydroxy-4'-methoxystilbene, as well as three novel compounds, yuccaols A, B, and C, with spiro-structures rarely occurring in the plant kingdom. It is suggested that yuccaols A-C are biosynthethized via attachment of a stilbenic derivative to the carbocationic intermediate of the oxidative flavanone-flavonol conversion.

Alfalfa (Medicago sativa L.) flavonoids. 1. Apigenin and luteolin glycosides from aerial parts.

Nine flavones and adenosine have been identified in aerial parts of alfalfa, and their structures were established by spectral (FABMS and NMR) techniques. Five of the identified compounds, including apigenin 7-O-[beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranosyl]-4'-O-beta-D-glucuronopyranoside, apigenin 7-O-[2-O-feruloyl-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranosyl]-4'-O-beta-D-glucuronopyranoside, apigenin 7-O-[2-O-feruloyl-[beta-D-glucuronopyranosyl(1-->3)]-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside], apigenin 7-O-[2-O-p-coumaroyl-[beta-D-glucuronopyranosyl(1-->3)]-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside], and luteolin 7-O-[2-O-feruloyl-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranosyl]-4'-O-beta-D-glucuronopyranoside, have not been reported before in the plant kingdom. Additionally, five known compounds, including apigenin 7-O-beta-D-glucuronopyranoside, apigenin 4'-O-beta-D-glucuronopyranoside, apigenin 7-O-[beta-D- glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside], luteolin 7-O-beta-D-glucuronopyranoside, and adenosine, were identified.

Saponins in alfalfa (Medicago sativa L.) root and their structural elucidation.

Twenty-four saponins have been identified in alfalfa roots, including 13 medicagenic acids, 2 zanhic acids, 4 hederagenins, 1 soyasapogenol A, 2 soyasapogenol B's, 1 soyasapogenol E, and 1 bayogenin glycoside. Ten of the identified compounds, including 3-O-[beta-D-glucopyranosyl(1-->3)-beta-D-glucopyranosyl]-28-O-beta-D- glucopyranoside medicagenate, 3-O-[alpha-L-rhamnopyranosyl(1-->2)-beta-D-glucopyranosyl(1-->2)-beta -D-glucopyranoside] medicagenic acid, 3-O-[beta-D-glucopyranosyl(1-->2)-beta-D-glucopyranosyl(1-->2)-beta-D -glucopyranosyl]-28-beta- D-glucopyranoside medicagenate, 3-O-[beta-D-glucuronopyranosyl methyl ester]-28-O-[beta-D-xylopyranosyl(1-->4)-alpha-L-rhamnopyranosyl(1--> 2)-alpha-L-arabinopyranoside] medicagenate, 3-O-[alpha-L-rhamnopyranosyl(1-->2)-beta-D-galactopyranosyl(1-->2)-be ta-D-glucuronopyranosyl]-21-O-alpha-L-rhamnopyranoside soyasapogenol A, 3-O-[beta-D-glucopyranosyl(1-->2)-beta-D-glucopyranosyl(1-->2)glucopy ranosyl]-28-O-[beta-D-xylopyranosyl(1-->4)-alpha-L-rhamnopyranosyl (1- ->2)-alpha-L-arabinopyranoside] medicagenate, 3-O-[beta-D-glucopyranosyl(1-->2)-beta-D-glucopyranosyl(1-->2)glucopy ranosyl]-28-O-¿beta-D-xylopyranosyl(1-->4)-)-[beta-D-apiofurano syl-(1 -->3)]- alpha-L-rhamnopyranosyl(1-->2)-alpha-L-arabinopyranoside¿ medicagenate, 3-O-[beta-D-glucopyranosyl(1-->2)-beta-D-glucopyranosyl(1-->2)-beta-D -glucopyranosyl]-28-O-[beta-D-xylopyranosyl(1-->4)-alpha-L-rhamnopyra nosyl(1-->2)-alpha-L-arabinopyranoside] zanhic acid, 3-O-[beta-D-glucopyranosyl(1-->2)-beta-D-glucopyranosyl(1-->2)-beta-D -glucopyranosyl]-28-O-¿beta-D-xylopyranosyl(1-->4)-[beta-D-apiofurano side-(1-->3)]- alpha-L-rhamnopyranosyl(1-->2)-alpha-L-arabinopyranoside¿zanhic acid, and 3-O-[beta-D-galactopyranosyl(1-->2)-beta-D-glucuronopyranosyl]-28- O-b eta-D-glucopyranoside bayogenin, were not reported before, and their structures were established by spectral (FAB-MS and NMR) techniques. In addition, 3-O-[alpha-L-rhamnopyranosyl(1-->2)-beta-D-galactopyranosyl(1-->2)-be ta-D-glucuronopyranoside] soyasapogenol E was identified in the roots for the first time.

Structure of steroidal saponins from underground parts of Allium nutans L.

Four steroidal glycosides including deltoside and nolinofuroside D and two novel saponins were isolated from underground parts of Allium nutans L. On the basis of the spectral (LSIMS and NMR) analysis, the structures of the new compounds were established as 25R Delta(5)-spirostan 3beta-ol-3-O-¿alpha-L-rhamnopyranosyl(1-->2)-[beta-D-glucopyranosyl(1 -->4)]-O-beta-D-galactopyranoside¿ and 25R Delta(5)-spirostan 1beta, 3beta-diol 1-O-beta-D-galactopyranoside. On the basis of the extraction efficiency, the concentration of saponins was established to be about 4% of dry matter, which makes this species a good source of steroidal saponins for commercial use.

Determination and toxicity of saponins from Amaranthus cruentus seeds.

The concentrations of four triterpene saponins present in amaranth seeds were determined with high-performance liquid chromatography. It was shown that the total concentration of saponins in seeds was 0. 09-0.1% of dry matter. In germinating seeds an increase in concentration to 0.18% was observed after 4 days of germination, which remained stable for the next 3 days and later dropped to 0.09%. Highly purified extracts from the seeds were tested for their toxicity against hamsters. The hydrophobic fraction obtained by the extraction of seeds with methylene chloride showed no toxicity; the behavior of tested animals was similar to that of the group given an equivalent dose of rapeseed oil. A crude saponin fraction, containing approximately 70% of pure saponins in the matrix, showed some toxicity; the approximate lethal dose was calculated as 1100 mg/kg of body weight. It is concluded that low contents of saponins in amaranth seeds and their relatively low toxicity guarantee that amaranth-derived products create no significant hazard for the consumer.

Effect of processing on the flavonoid content in buckwheat (Fagopyrum esculentum Möench) grain.

Six flavonoids have been isolated and identified in buckwheat grain. These are rutin, orientin, vitexin, quercetin, isovitexin, and isoorientin. Rutin and isovitexin are the only flavonoid components of buckwheat seeds while hulls contain all six identified compounds. The total flavonoid concentration in the seeds was 18.8 and in the hulls 74 mg/100 g of dry matter. Dehulling the grain by using different temperature regimes resulted in drastic reductions of the total flavonoid concentration in the grain (by 75% of the control) and smaller but significant (15-20%) reduction in the hulls.

Isolation and studies of the mutagenic activity of saponins in the Ames test.

Mutagenic activity of medicagenic acid, medicagenic acid 3-0-glucopyranoside and soyasaponin I was tested by the Ames method with S. typhimurium strains TA97, TA98, TA100, TA102 in the absence and the presence of metabolic activation (S9 mix). These saponins have been isolated and identified from alfalfa roots and clover Trifolium incarnatum seeds. All of them were found to be non-toxic and non-mutagenic for testing doses.