The aim of the study was to determine phytoestrogen content in fresh-cut legume forage. This issue has been much discussed in recent years in connection with the health and safety of feedstuffs and thus livestock health. The experiments were carried out on two experimental plots at Troubsko and Vatín, Czech Republic during June and July in 2015. Samples were collected of the four forage legume species perennial red clover (variety "Amos"), alfalfa (variety "Holyně"), and annuals Persian clover and Alexandrian clover. Forage was sampled twice at regular three to four day intervals leading up to harvest and a third time on the day of harvest. Fresh and wilted material was analyzed using liquid chromatography-mass spectrometry (LC-MS). Higher levels ( p < 0.05) of isoflavones biochanin A (3.697 mg·g (-1) of dry weight) and formononetin (4.315 mg·g (-1) of dry weight) were found in red clover than in other species. The highest isoflavone content was detected in red clover, reaching 1.001% of dry matter ( p < 0.05), representing a risk for occurrence of reproduction problems and inhibited secretion of animal estrogen. The phytoestrogen content was particularly increased in wilted forage. Significant isoflavone reduction was observed over three to four day intervals leading up to harvest.

Department of Animal Nutrition and Forage Production Faculty of Agronomy Mendel University in Brno Zemědělsk and 225; 1 1665 Brno CZ 613 00 Czech Republic

Department of Chemistry and Biochemistry Faculty of Agronomy Mendel University in Brno Zemědělsk and 225; 1 1665 Brno CZ 613 00 Czech Republic

Research Institute for Fodder Crops Ltd Troubsko Zahradn and 237; 1 Troubsko 664 41 Czech Republic

Zobrazit více v PubMed

Kašparová M. Phytoestrogens from red clover. Prakt. Lék. 2013;9:201–203.

Opletal L., Šimerda B. Přírodní Látky a Jejich Biologická Aktivita. 4. Fytoestrogeny Přírodního Původu, Výskyt v Krmivovém (Potravním) Řetězci, Pozitivní a Negativní Účinky. Studie Vědeckého Výboru Výživy Zvířat při Mze. Výzkumný Ústav Živočišné Výroby Praha-Uhříněves; Prague, Czech Republic: 2010.

Lincová D., Farghali D. Základní a Aplikovaná Farmakologie. 2nd ed. Galén; Prague, Czech Republic: 2007.

Chen Y., Huang C., Zhou T., Zhang S., Chen G. Biochanin A induction of sulfotransferases in rats. J. Biochem. Mol. Toxicol. 2010;24:102–114. doi: 10.1002/jbt.20318. PubMed DOI

De Lucas A., Panter S., Mouradov A., Rochfort S., Smith K.F., Spangenberg G. Assessment of nutritional characteristics of virus-resistant transgenic white clover (Trifolium repens L.) grown under field and glasshouse conditions. Mol. Breed. 2015;35:147. doi: 10.1007/s11032-015-0341-2. DOI

Kalač P., Míka V. Přirozené Škodlivé Látky v Rostlinných Krmivech. 1st ed. Ústav Zemědělských a Potravinářských Informací; Prague, Czech Republic: 1997.

Kwiatkowska E. Phytoestrogens in the prevention of diseases. Postępy Fitoter. 2009;2:107–112.

Ramírez-Restrepo C.A., Barry T.N. Alternative temperate forages containing secondary compounds for improving sustainable productivity in grazing ruminants. Anim. Feed Sci. Technol. 2005;120:179–201. doi: 10.1016/j.anifeedsci.2005.01.015. DOI

Český Hydrometeorologický Ústav. [(accessed on 2 December 2015)]. Available online: http://portal.chmi.cz.

Xiao Y., Zhang J., Jia T.T., Pang X.P., Guo Z.G. Effects of alternate furrow irrigation on the biomass and quality of alfalfa (Medicago sativa) Agric. Water Manag. 2015;161:147–154. doi: 10.1016/j.agwat.2015.07.018. DOI

Spagnuolo P., Rasini E., Luini A., Legnaro M., Luzzani M., Casareto E., Cosentino M. Isoflavone content and estrogenic activity of different batches of red clover (Trifolium pratense L.) extracts: An in vitro study in MCF-7 cells. Fitoterapia. 2014;94:62–69. doi: 10.1016/j.fitote.2014.01.027. PubMed DOI

Graves M.E., McLean N., Jones G., Martin R.C. Pasture and sheep performance response to sod-seeding red clover (Trifolium pratense L.) or white clover (Trifolium repens L.) into naturalized pastures in eastern Canada. Anim. Feed Sci. Technol. 2012;177:7–14.

El-Lithy M., El-Batanony N., Moreno S., Bedmar E.A. Selected rhizobial strain isolated from wild-grown Medicago monspeliaca improves productivity of non-specific host Trifolium alexandrinum. Appl. Soil Ecol. 2014;73:134–139. doi: 10.1016/j.apsoil.2013.08.013. DOI

Simonet A.M., Stochmal A., Oleszek W., Macías F.A. Saponins and polar compounds from Trifolium resupinatum. Phytochemistry. 1999;51:1065–1067. doi: 10.1016/S0031-9422(99)00192-2. DOI

Klejdus B., Lojková L., Plaza M., Snóblová M., Stěrbová D. Hyphenated technique for the extraction and determination of isoflavones in algae: Ultrasound-assisted supercritical fluid extraction followed by fast chromatography with tandem mass spectrometry. J. Chromatogr. A. 2010;1217:7956–7965. doi: 10.1016/j.chroma.2010.07.020. PubMed DOI

Wu Q., Wang M., Simon J.E. Determination of isoflavones in red clover and related species by high-performance liquid chromatography combined with ultraviolet and mass spectrometric detection. J. Chromatogr. A. 2003;24:195–209. doi: 10.1016/j.chroma.2003.08.001. PubMed DOI

Barreira J.C.M., Visnevschi-Necrasov T., Nunes E., Cunha S.C., Pereira G., Oliveira M.B.P.P. Medicago spp. as potential sources of bioactive isoflavones: Characterization according to phylogenetic and phenologic factors. Phytochemistry. 2015;116:230–238. doi: 10.1016/j.phytochem.2015.04.011. PubMed DOI

Nykänen-Kurki P., Saloniemi H., Kallela K., Saastamoinen I. Phyto-oestrogen content and oestrogenic effect of white clover. In: Frame J., editor. White Clover in Europe: State of the Art. Food and Agriculture Organization of the United Nations. Food and Agriculture Organization of the United Nations; Rome, Italy: 1993. [(accessed on 1 December 2015)]. Available online: http://www.fao.org/docrep/v2350e/v2350e0e.htm.

De Rijke E., Zafra-Gomez A., Ariese F., Brinkman U.A.T., Gooijer C. Determination of isoflavone glucoside malonates in Trifolium pratense L. (red clover) extracts: Quantification and stability studies. J. Chromatogr. A. 2001;932:55–64. doi: 10.1016/S0021-9673(01)01231-6. PubMed DOI