The aim of this study was to determine the degradation of dietary isoflavones in rumen fluid under 2 feeding regimens. The experiments were performed in vitro using a rumen fluid buffer system. The rumen fluid was taken from cows fed either a hay diet or a concentrate-rich diet (the diet consisted of 34.6% maize silage, 17.6% haylage, 12.8% alfalfa hay, and 35.0% supplemental mixture on a dry matter basis). As a source of isoflavones, 40% soybean extract (Biomedica, Prague, Czech Republic) at levels of 5, 25, 50, and 75 mg per 40 mL of rumen fluid was used. Samples of soybean extract were incubated in triplicate at 39°C for 0, 3.0, 6.0, 12.0, and 24.0 h in incubation solution. The metabolism of daidzein and genistein was faster under concentrate-rich diet conditions. In general, production of equol started after 3 to 6 h of incubation and reached the highest rate after approximately 12 h of incubation regardless of the type of diet or concentration of extract. In most of the experiments, production of equol continued after 24 h of incubation. Generally, equol production was greater under the hay diet conditions. Furthermore, experiments with higher amounts of added soybean extract revealed possible inhibitory effects of high levels of isoflavones on the rumen microflora.
- MeSH
- bachor metabolismus MeSH
- dieta MeSH
- fyziologie výživy zvířat fyziologie MeSH
- isoflavony aplikace a dávkování analýza metabolismus MeSH
- krmivo pro zvířata MeSH
- laktace MeSH
- siláž MeSH
- skot MeSH
- zvířata MeSH
- Check Tag
- skot MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Compounds with estrogenic potencies and their adverse effects in surface waters have received much attention. Both anthropogenic and natural compounds contribute to overall estrogenic activity in freshwaters. Recently, estrogenic potencies were also found to be associated with cyanobacteria and their blooms in surface waters. The present study developed and compared the solid phase extraction and LC-MS/MS analytical approaches for determination of phytoestrogens (8 flavonoids - biochanin A, coumestrol, daidzein, equol, formononetin, genistein, naringenin, apigenin - and 5 sterols - ergosterol, β-sitosterol, stigmasterol, campesterol, brassicasterol) and cholesterol in water. The method was used for analyses of samples collected in stagnant water bodies dominated by different cyanobacterial species. Concentrations of individual flavonoids ranged from below the limit of detection to 3.58 ng/L. Sterols were present in higher amounts up to 2.25 μg/L. Biological potencies of these phytoestrogens in vitro were characterized using the hERα-HeLa-9903 cell line. The relative estrogenic potencies (compared to model estrogen - 17β-estradiol) of flavonoids ranged from 2.25E-05 to 1.26E-03 with coumestrol being the most potent. None of the sterols elicited estrogenic response in the used bioassay. Estrogenic activity was detected in collected field water samples (maximum effect corresponding to 2.07 ng/L of 17β-estradiol equivalents, transcriptional assay). At maximum phytoestrogens accounted for only 1.56 pg/L of 17β-estradiol equivalents, contributing maximally 8.5% of the total estrogenicity of the water samples. Other compounds therefore, most likely of anthropogenic origin such as steroid estrogens, are probably the major drivers of total estrogenic effects in these surface waters.
- MeSH
- chemické látky znečišťující vodu analýza MeSH
- cholestadienoly MeSH
- cholesterol analogy a deriváty MeSH
- estradiol analýza MeSH
- estrogeny analýza MeSH
- estron analýza MeSH
- fytoestrogeny analýza MeSH
- fytosteroly MeSH
- genistein analýza MeSH
- HeLa buňky MeSH
- isoflavony analýza MeSH
- lidé MeSH
- receptory pro estrogeny metabolismus MeSH
- sinice účinky léků metabolismus MeSH
- sitosteroly analýza MeSH
- sladká voda MeSH
- steroly analýza MeSH
- tandemová hmotnostní spektrometrie MeSH
- voda MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Soybean-derived isoflavones belong to the family of biologically active phytoestrogens. The purpose of this study was to develop a sensitive method, which permits quantification of the soybean isoflavonoids and equol in bovine rumen fluid and milk using LC-MS-(TOF). The samples of rumen fluid and milk were obtained from 12 lactating dairy cows ingesting 7,500-9,500 mg of total isoflavones daily. The validation of the developed method showed the limits of quantification to be in the range of 0.9-5.0 ng/mL. The precision was determined as relative standard deviation, which was lower than 25% in all cases. The recoveries of the most isoflavonoids were satisfactory. Lower recoveries of daidzin and glycitin can be solved by adding an internal standard. The presented method will be useful for kinetic studies of isoflavone metabolism in ruminants due to simultaneous quantification of free aglycones and glycosides in the rumen fluid.
- MeSH
- bachor MeSH
- chemické techniky analytické metody MeSH
- chromatografie kapalinová * MeSH
- gastrointestinální obsah chemie MeSH
- Glycine max chemie metabolismus MeSH
- isoflavony analýza MeSH
- kinetika MeSH
- mléko chemie MeSH
- skot MeSH
- tandemová hmotnostní spektrometrie * MeSH
- zvířata MeSH
- Check Tag
- skot MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
This review discusses the potential contribution of phytoestrogens and mycoestrogens to in vitro estrogenic activities occurring in surface waters and in vivo estrogenic effects in fish. Main types, sources, and pathways of entry into aquatic environment of these detected compounds were summarized. Reviewed concentrations of phyto/mycoestrogens in surface waters were mostly undetectable or in low ng/L ranges, but exceeded tens of μg/L for the flavonoids biochanin A, daidzein and genistein at some sites. While a few phytosterols were reported to occur at relatively high concentrations in surface waters, information about their potencies in in vitro systems is very limited, and contradictory in some cases. The relative estrogenic activities of compounds (compared to standard estrogen 17β-estradiol) by various in vitro assays were included, and found to differ by orders of magnitude. These potencies were used to estimate total potential estrogenic activities based on chemical analyses of phyto/mycoestrogens. In vivo effective concentrations of waterborne phyto/mycoestrogens were available only for biochanin A, daidzein, formononetin, genistein, equol, sitosterol, and zearalenone. The lowest observable effect concentrations in vivo were reported for the mycoestrogen zearalenone. This compound and especially its metabolites also elicited the highest in vitro estrogenic potencies. Despite the limited information available, the review documents low contribution of phyto/mycoestrogens to estrogenic activity in vast majority of surface waters, but significant contribution to in vitro responses and potentially also to in vivo effects in areas with high concentrations.
- MeSH
- chemické látky znečišťující vodu * analýza toxicita MeSH
- fytoestrogeny * analýza toxicita MeSH
- isoflavony * analýza toxicita MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
The increased content of isoflavonoids in dry cell suspension and nutrient medium was observed after application of electric current and AgNO3 on Genista tinctoria L. cultures in vitro. The highest content of genistin (1.7 mg g(- 1) DW - dry weight) was measured in the dry cell suspension culture after 30 min elicitation of 10 V and 6 h cultivation and daidzein content (3.5 mg g(- 1) DW) was measured after 60 min elicitation of 5 V and 24 h cultivation. In the case of AgNO3 elicitation, the content of genistin in dry cell suspension culture (0.5 mg g(- 1) DW) was highest after 48 h of AgNO3 treatment and concentration of 5.9 × 10(- 4) mol/L. The AgNO3 concentration of 5.9 × 10(- 4) mol/L was also the most effective combination for daidzein production (0.9 mg g(- 1) DW) after 168 h. The results of this study show that the secondary metabolites could also be released from G. tinctoria L. cells into the nutrient medium.
BACKGROUND: Isoflavones and coumestrol from dietary legumes are plant constituents showing multiple beneficial effects on humans. Owing to their ability to bind with mammalian estrogenic receptors and thereby intervention in several kinds of hormone-related cancers, they have received much attention. Soybean (Glycine max) is currently the major source of isoflavonoids in human diet. However, dozens of tropical and subtropical leguminous species remain unexplored for their isoflavonoids content. RESULTS: We have analyzed 55 extracts from 41 tropical and subtropical legume species used either in human or animal diet by high-performance liquid chromatography for the content of soy isoflavones, biochanin A, daidzein, daidzin, formononetin, genistein, genistin, sissotrin, ononin and the coumestan coumestrol. Genistein and biochanin A were the most abundant compounds. The highest content of genistein was found in aerial parts of Andira macrothyrsa, seeds of Pachyrhizus tuberosus and aerial parts of Calopogonium mucunoides (598, 250 and 184 µg g(-1), respectively) and biochanin A in aerial parts of Cratylia argentea, C. mucunoides and flowers of A. macrothyrsa (76, 53 and 40 µg g(-1), respectively). CONCLUSION: None of the samples tested was richer overall source of soy isoflavones and coumestrol than soybean; nevertheless several species (C. mucunoides or A. macrothyrsa) may serve as a promising source of individual compounds.
- MeSH
- dieta * MeSH
- Fabaceae chemie MeSH
- genistein analýza MeSH
- Glycine max chemie MeSH
- isoflavony analýza MeSH
- kumestrol analýza MeSH
- lidé MeSH
- nadzemní části rostlin MeSH
- podpora zdraví MeSH
- rostlinné extrakty chemie MeSH
- semena rostlinná chemie MeSH
- tropické klima MeSH
- vysokoúčinná kapalinová chromatografie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
A direct analysis in real-time (DART) ion source coupled to a high-resolution orbitrap mass spectrometer was used for the quantitative analysis of isoflavones isolated from soybeans. For the isolation of genistein, daidzein, glycitein, and their respective acetyl, malonyl, and glucoside forms, an extraction employing 80% aqueous MeOH enhanced by sonication was used. As far as the total isoflavones (expressed as aglycones) were to be determined, an acid hydrolysis with 80% aqueous EtOH and refluxing had to be employed, while in the latter case a good agreement of the results with the data generated by the UHPLC-orbitrap MS method was achieved, in the case of the analysis of non-hydrolyzed extracts, some overestimation of the results as compared with those generated by UHPLC-orbitrap MS was observed. A careful investigation of this phenomenon showed that the free aglycones originated from the conjugated forms of isoflavones in the DART ion source, thus contributing significantly to the "free" genistein/daidzein/glycitein signals during the DART analysis. Good recoveries (95-102%) and repeatabilities (RSD: 7-15%) were obtained at the spiking levels of 0.5, 1, and 0.05 g/kg, for daidzein, genistein, and glycitein, respectively. The limits of detection estimated for the respective analytes were 5 mg/kg.
Kručinka barvířská (Genista tinctoria L.) – obsahové látky a biologická aktivita Cílem práce bylo uvést všechny důležité obsahové látky v rostlině Genista tinctoria, které vykazují různou biologickou aktivitu. Jsou zde také zmíněny metody chemického hodnocení těchto látek.
The paper aims to specify all important active principles in the herb Genista tinctoria which exert different biological activities. It also mentions the methods of chemical evaluation of these substances.
- Klíčová slova
- in vitro, quinolizidine alkaloids, isoflavonoids,
- MeSH
- alkaloidy analýza farmakologie MeSH
- Genista chemie MeSH
- isoflavony analýza farmakologie MeSH
- léčivé rostliny chemie MeSH
New hyphenated technique for the extraction and determination of isoflavones in sea and freshwater algae and cyanobacteria was developed. The method consists of sonication sample pretreatment, extraction by supercritical CO(2) modified by 3% (v/v) of MeOH/H(2)O mixture (9:1, v/v) at 35 MPa and 40°C for 60 min, fast chromatography analysis by the means of Agilent 1200 Series Rapid Resolution and MS/MS determination. Agilent 1200 Series RRLC was used with Zorbax SB-CN chromatographic column (100 mm × 2.1mm, particle size 3.5 μm), 3μl injection volume, mobile phase consisting of 0.2% (v/v) acetic acid in water (solvent A) and acetonitrile (solvent B) and used with linear gradient (30% B at 0 min, from 0 min to 3 min up to 50% B, from 3 to 6 min up to 80% B and from 6 to 10 min down to 30% B). The flow-rate was 0.4 mL/min, column oven temperature 35°C. MS detector Agilent Technologies 6460 Triple quadrupole LC/MS with Agilent Jet Stream was used in a negative ESI mode under following conditions: gas temperature 350°C, gas flow 13 L/min, nebulizer gas pressure 50 psi, sheath gas temperature 400°C, sheath gas flow 12L/min, capillary voltage was 4 kV. Samples were analysed in the multiple reaction monitoring (MRM) mode. Eight isoflavone compounds were found for the first time in seven real samples of sea algae and in three control samples of freshwater algae and cyanobacteria. Usual optimisation study of extraction parameters was performed. Pressure and temperature optima for algae matrix are different from those obtained sooner for other matrices for most of the analytes, but the results of modifier optimisation study are in good accordance with those obtained sooner for spiked samples and red clover matrix. It seems that matrix has very small or no effect on the modifier selection. Two different approaches of sonication pretreatment were tested: sonication bath and the thorn instrument. In longer extraction time experiments, thorn sonication was more efficient and recovery of following supercritical fluid extraction was higher.
- MeSH
- chromatografie kapalinová metody MeSH
- isoflavony analýza izolace a purifikace MeSH
- Phaeophyceae chemie MeSH
- Rhodophyta chemie MeSH
- superkritická fluidní chromatografie přístrojové vybavení metody MeSH
- tandemová hmotnostní spektrometrie metody MeSH
- ultrazvuk MeSH
- Publikační typ
- časopisecké články MeSH
- hodnotící studie MeSH
- práce podpořená grantem MeSH
The antioxidant properties of pomiferin, isopomiferin, osajin and catalposide are evaluated. The electrochemical behaviour of these compounds at a carbon paste electrode was studied using square wave voltammetry. Oxidative signals, optimized frequencies and appropriate pH acetate buffer conditions were determined. The detection limits (3S/N) for pomiferin, isopomiferin, osajin and catalposide were estimated to be 50 pg/ml, 800 pg/ml, 40 pg/ml and 10 ng/ml, respectively. Furthermore, spectrometric test was employed with 2,2-diphenyl-1-picrylhydrazyle (DPPH) to evaluate the antioxidant activities of these compounds. Based on the obtained results, the highest antioxidant activity measured by DPPH tests was found at pomiferin followed by isopomiferin. The activities of osajin and catalposide were undetectable. The protective effects of pomiferin, isopomiferin, osajin and catalposide on DNA exposed to oxygen radicals in vitro were also studied. Changes in height of oxidative signals for the four bases (guanine, thymine, adenine and cytosine) were measured for DNA exposed to oxygen radicals, generated by Fenton's reaction, non-oxidized ssDNA (50 microg/ml) displayed well developed signals; however, after oxidative damage the observed oxidative signals decreased. Significant protective effects were observed for pomiferin and osajin. Decreased effect was observed for isopomiferin while a further reduced protective effect was seen for DNA exposed to catalposide. Based on the obtained results, pomiferin had the highest antioxidant activity followed by isopomiferin, osajin and catalposide.
- MeSH
- antioxidancia analýza farmakologie chemie MeSH
- benzopyrany MeSH
- elektrochemie MeSH
- financování organizované MeSH
- glukosidy MeSH
- isoflavony analýza farmakologie chemie MeSH
- koncentrace vodíkových iontů MeSH
- molekulární struktura MeSH
- oxidace-redukce účinky léků MeSH
- spektrofotometrie metody MeSH
- Publikační typ
- srovnávací studie MeSH
