Means and methods for reducing estrogenic effect of xenoestrogen

Abstract

The present invention relates to a composition comprising a compound having formula (I), preferably selected from (E)-2,4-dihydroxy-6-(10-hydroxy-6-oxo-1-undecen-1-yl)benzoic acid or (E)-1-(3,5-dihydroxy-phenyl)-10-hydroxy-1-undecen-6-one, a method for reducing estrogenic effect(s) using said compound, use of said compound, and a composition comprising said compound for use in treatment, amelioration and / or prevention of symptoms associated with estrogenic effect(s) of a xenoestrogen.

Description

MEANS AND METHODS FOR REDUCING ESTROGENIC EFFECT OF XENOESTROGEN

[0001] The present invention relates to a composition, a method for reducing estrogenic effect(s), use of a compound and a composition for use in treatment, amelioration and / or prevention.

[0002] Xenoestrogens are compounds capable of eliciting a hormonal, in particular an estrogenic response in an animal body, which is different from their place of origin or synthesis. Xenoestrogens may be of synthetic origin, or stem from natural organisms such as plants or fungi.

[0003] In some cases, an effect on the endocrine system may be desired, e.g. in xenoestrogen- effected contraception. In other cases however, such estrogenic effects are associated with increased cancer risk, reduced fertility in males and females, environmental damages etc.

[0004] Zearalenone and structurally and functionally similar derivatives thereof, such as e.g. zearalenol, are produced by various Fusarium species, and belong to the group of fungal xenoestrogens, also referred to as mycoestrogens. Such mycoestrogens are known to cause serious damages in the agriculture industry in terms of both, animal health as well as economically. In particular, zearalenone has been described to negatively affect male and female animals, causing damage to liver, hematotoxic effects, and, most prominently, disorders associated with estrogenic effect such as infertility, vulva swelling, uterus enlargement, pseudopregnancy, stillbirths, ovary cysts, enlarged mammary glands, feminization of testicular atrophy, reduced sperm concentration etc. (Ropejko et al. 2021. Toxins 13(1): 25).

[0005] Phytoestrogens are produced by plants, and comprise various compounds of the groups of coumestans, prenylflavonoids, lignans, stilbenes and isoflavones. Numerous studies have shown adverse effects of phytoestrogens on animals' reproduction organs and functions, e.g. Rochester et al. 2009. Comp Biochem Physiol A Mol Integr Physiol 154(3):279; Grgic et al. 2021 . Toxins 13(12):836, associated with the estrogenic effect of these compounds.

[0006] Notwithstanding the above, isoflavones have also gained attention for beneficial health effects, for instance in individuals suffering from obesity, metabolic syndrome, type 2 diabetes, osteoporosis, and / or irritable bowel syndrome, for providing protection against reactive oxygen species, and potentially supporting neuronal health and cognitive performance (e.g. Rietjens et al. 2016. Br J Pharmacol 174(11):1263).

[0007] While most of the undesirable effects can be directly linked to the estrogenic activity of xenoestrogens, in particular of phytoestrogens, the mode-of-action for many of the beneficial effects is either unknown or assumed to relate to a non-estrogen related mechanism, e.g. CpG hypermethylation, AMP-activated protein kinase activation, kinase inhibition,antioxidant / electrophile response element induction, etc. (Rietjens et al. 2016. Br J Pharmacol 174(11):1263). Effects attributable to estrogenic effects of xenoestrogens on the other hand, are caused by the binding of the xenoestrogens to estrogen receptor (ER) isoforms alpha (ERa) and / or beta (ERP).

[0008] In view of the prior art as outlined above, it is an objective of the present invention to provide means and methods that allow individuals to benefit from the desirable effects of xenoestrogens, such as isoflavones, without their unfavorable estrogenic effect.

[0009] This objective is achieved by a composition (e.g. a nutritional composition) comprising at least one isoflavone, and at least a second compound having formula (I):

[0010] wherein R is selected from H and COOH.

[0011] Such a composition allows providing the desirable non-estrogenic effects of the at least one isoflavone, but has a reduced estrogenic potential. In other words, owing to the presence of the at least second compound, such a composition elicits reduced estrogenic effect or even no estrogenic effects at all, e.g. upon consumption.

[0012] In some embodiments, the composition further comprises one or more further compounds selected from: One or more chemical(s) capable of modifying mycotoxin(s); one or more polypeptide(s) capable of modifying mycotoxin(s); one or more organic absorbent(s); one or more inorganic absorbent(s); one or more live, inactivated, lyophilized and / or dormant microorganism(s) capable of modifying mycotoxin(s); one or more plant product(s); one or more flavoring compound(s); and one or more vitamin(s). In particular, said modifying mycotoxin(s) is a detoxification of said mycotoxin(s); or a reduction in toxicity of said mycotoxin(s).

[0013] In some embodiments, said one or more chemicals capable of modifying mycotoxin(s) is / are one or more metabisulfite salt(s), such as sodium metabisulfite.

[0014] In some embodiments, said one or more polypeptide(s) capable of modifying mycotoxin(s) is / are one or more peptidase(s), hydrolase(s), esterase(s), lactonase(s), epoxidase(s), peroxidase(s), and / or peroxygenase(s). Preferably, said one or more further polypeptide(s) capable of modifying one or more mycotoxin(s) is / are e.g. fumonisin esterase (e.g. as disclosed in WO 2016 / 134387 A1), ochratoxin peptidase (e.g. as disclosed in WO 2023 / 025938 A1 , or inWO 2022 / 129517 A2), zearalenone lactonase (e.g. as disclosed in WO 2020 / 025580 A1 , or in WO 2022 / 073649 A1), ergopeptine hydrolase (e.g. as disclosed in WO 2014 / 056006 A1).

[0015] In some embodiments, said one or more organic absorbent(s) is / are live, inactivated, lyophilized, dormant, and / or dead whole-yeast or yeast-derived product such as e.g. yeast cell wall, or yeast oligosaccharides such as e.g. mannan.

[0016] In some embodiments, said one or more inorganic absorbent(s) is / are diatomaceous earth and / or clay mineral such as e.g. kaolins or kaolinites, smectites such as e.g. montmorillonites, illites or chlorites; in particular bentonite.

[0017] In some embodiments, said one or more live, inactivated, lyophilized and / or dormant microorganism(s) capable of modifying mycotoxin(s) is / are selected from the group consisting of Trichosporon and Apiotrichum genera (e.g. as disclosed in WO 03 / 053161 A1) and the Coriobacteriaceae family (e.g. as disclosed in EP 3 501 526 A1).

[0018] In some embodiments, said one or more plant product(s) is / are e.g., seaweed, preferably seaweed meal; and / or algae, preferably algae meal; and / or thistle, preferably thistle seeds; and / or glycyrrhiza plant preparation, preferably glycyrrhiza meal and / or glycyrrhiza extract, e.g. as disclosed in WO 2018 / 121881 A1.

[0019] In some embodiments, said one or more flavoring compound(s) is / are e.g., plant extract e.g. from oregano, thyme, Wintergreen, caraway, marjoram, mint, peppermint, anise, orange, lemon, fennel, star anise, clove, cinnamon and / or garlic; and / or essential oil such as e.g. D- limonene, y-terpinene, p-cymene, 2-carene, linalool oxide, isomenthone, camphor, linalool, terpinen-4-ol, 2-isopropyl-1-methoxy-4-methylbenzene, L-menthol, ethylamine, a-terpineol, - caryophyllene, D-carvone, methyl salicylate, a-caryophyllene, lavandulyl acetate, caryophyllene oxide, eugenol, thymol and / or carvacrol.

[0020] In some embodiments, said one or more vitamin(s) is / are e.g. vitamin A, D, E, K, C, B1 , B2, B3, B4, B5, B6, B7, B8, B9 and / or B12.

[0021] In some embodiments of the composition according to the invention, the at least one isoflavone is selected from genistein (CAS no. 446-72-0), daidzein (CAS no. 486-66-8), glycitein (CAS no. 40957-83-3) and equol (CAS no. 531-95-3).

[0022] In some embodiments of the composition according to the invention, (i) the composition is foodstuff; fodder; feed; silage; wet distillers grain; dried distillers grain with solubles; intermediate / s thereof; and / or mixture / s thereof; and / or (ii) the composition comprises cereal(s) such as rice, wheat, rye, oats, corn or maize, barley, sorghum, and / or soy or other Fabaceae plant(s) suitable for human and / or animal consumption; and / or product(s) thereof.

[0023] In another aspect, the invention relates to a method for reducing estrogenic effect(s) of a xenoestrogen, comprising forming a mixture of the xenoestrogen with a compound having formula (I):

[0024] wherein R is selected from H and COOH.

[0025] In particular, said step of forming a mixture of the xenoestrogen with the compound, comprises contacting the xenoestrogen with the compound.

[0026] In some embodiments, the invention relates to a method for reducing estrogenic effect(s) of a composition (e.g. a nutritional composition) comprising a xenoestrogen, comprising forming a mixture of the composition comprising the xenoestrogen with said compound. In particular, said step of forming a mixture of the composition with the compound, comprises contacting the composition with the compound.

[0027] Estrogenic effect(s) may be determined e.g. by using the human endometrial adenocarcinoma cell line “Ishikawa” as described e.g. by Grgic et al. 2022. Arch Toxicol. 96(12): 3385-3402.

[0028] In a further aspect, the invention relates to a method for reducing estrogenic effect(s) of a xenoestrogen on an animal cell, comprising the step of contacting the animal cell with a compound having formula (I):(0 /

[0029] wherein R is selected from H and COOH.

[0030] In particular, said animal cell (human or non-human animal cell) is an animal cell comprising one or more estrogen receptor(s), preferably one or more of the estrogen receptor isoform(s) alpha and beta.

[0031] In some embodiments of the invention, said compound having formula (I) is provided to the animal cell prior to a contact with the xenoestrogen. In some embodiments, said compound having formula (I) is provided to the animal cell at the same time as the xenoestrogen. In some embodiments, said compound having formula (I) is provided to the animal cell after a previous contact of the xenoestrogen with the animal cell.

[0032] In another aspect, the invention relates to a method for manufacturing a composition (e.g. for manufacturing a nutritional composition), the method comprising the steps of providing a compound having formula (I):(0 /

[0033] wherein R is selected from H and COOH; and contacting the composition with the compound.

[0034] Hereby, a composition will be produced which is less prone to causing estrogenic effect(s). In other words, a such produced composition has a reduced estrogenic potential. Thus, such a method for manufacturing a composition is a method for increasing the safety of said composition.

[0035] In some embodiments of the method for manufacturing a composition according to the invention, (i) the composition is foodstuff; fodder; feed; silage; wet distillers grain; dried distillers grain with solubles; intermediate / s thereof; and / or mixture / s thereof; and / or (ii) the composition comprises cereal(s) such as rice, wheat, rye, oats, corn or maize, barley, sorghum, and / or soy or other Fabaceae plant(s) suitable for human and / or animal consumption; and / or product(s) thereof.

[0036] In some embodiments, the composition of the method for manufacturing a composition comprises a xenoestrogen.

[0037] In some embodiments of a method according to the invention, the xenoestrogen is a phytoestrogen, preferably an isoflavone, more preferably an isoflavone selected from genistein, daidzein, glycitein and equol.

[0038] In a further aspect, the invention relates to a use of a compound having formula (I):(I),

[0039] wherein R is selected from H and COOH; for reducing estrogenic effect(s) of a xenoestrogen.

[0040] In some embodiments of the use according to the invention, the xenoestrogen is a phytoestrogen, preferably an isoflavone, more preferably an isoflavone selected from genistein, daidzein, glycitein and equol.

[0041] In a further aspect, the invention relates to a composition (e.g. nutritional composition) for use in treatment, amelioration and / or prevention of symptoms associated with estrogenic effect(s) of a xenoestrogen; wherein the composition comprises at least a compound having formula (I):(0 /

[0042] wherein R is selected from H and COOH.

[0043] In some embodiments, the composition (e.g. nutritional composition) for use in treatment, amelioration and / or prevention of symptoms associated with estrogenic effect(s) of a xenoestrogen, (i) is foodstuff; fodder; feed; silage; wet distillers grain; dried distillers grain with solubles; intermediate / s thereof; and / or mixture / s thereof; and / or (ii) comprises cereal(s) such as rice, wheat, rye, oats, corn or maize, barley, sorghum, and / or soy or other Fabaceae plant(s) suitable for human and / or animal consumption; and / or product(s) thereof.

[0044] In some embodiments of the composition (e.g. nutritional composition) for use according to the invention, the xenoestrogen is a phytoestrogen, preferably an isoflavone, more preferably an isoflavone selected from genistein, daidzein, glycitein and equol.

[0045] In some embodiments, the compound having formula (I) is (E)-2,4-dihydroxy-6-(10- hydroxy-6-oxo-1-undecen-1-yl)benzoic acid or (E)-1-(3,5-dihydroxy-phenyl)-10-hydroxy-1- undecen-6-one, as described e.g. by Vekiru et al. 2016. World Mycotoxin Journal 9(3): 353-363. In some embodiments, the compound is a mixture of both, (E)-2,4-dihydroxy-6-(10-hydroxy-6- oxo-1 -undecen-1-yl)benzoic acid and (E)-1-(3,5-dihydroxy-phenyl)-10-hydroxy-1-undecen-6-one. In some embodiments, the compound is provided by modifying (3S,11E)-14,16-dihydroxy- 3-methyl-3,4,5,6,9,10-hexahydro-1 H-2-benzoxacyclotetradecin-1 ,7(8H)-dione to (E)-2,4- dihydroxy-6-(10-hydroxy-6-oxo-1-undecen-1-yl)benzoic acid and / or (E)-1-(3,5-dihydroxy- phenyl)-10-hydroxy-1-undecen-6-one; e.g. by enzymatically modifying (3S,11E)-14,16- dihydroxy-3-methyl-3,4,5,6,9,10-hexahydro-1 H-2-benzoxacyclotetradecin-1 ,7(8H)-dione to form (E)-2,4-dihydroxy-6-(10-hydroxy-6-oxo-1-undecen-1-yl)benzoic acid and / or (E)-1-(3,5-dihydroxy- phenyl)-10-hydroxy-1-undecen-6-one.

[0046] The invention is further characterized by the following items:

[0047] Item 1. Composition (e.g. nutritional composition) comprising at least a first compound, wherein the at least first compound is at least one isoflavone; and at least a second compound having formula (I):(I), preferably wherein R is selected from H and COOH.

[0048] Item 2. Composition (e.g. nutritional composition) comprising at least a first compound, wherein the at least first compound is at least one isoflavone; and at least a second compound having formula (I):(I), preferably wherein R is selected from H and COOH; wherein an estrogenic potential of the composition is reduced compared to a composition not comprising the at least second compound.

[0049] Item 3. Composition (e.g. nutritional composition) comprising at least one isoflavone;(3S,11 E)-14,16-dihydroxy-3-methyl-3,4,5,6,9,10-hexahydro-1 H-2-benzoxacyclotetradecin-1 ,7(8H)-dione; and means for converting (3S,11E)-14,16-dihydroxy-3-methyl-3,4,5,6,9,10- hexahydro-1 H-2-benzoxacyclotetradecin-1,7(8H)-dione to a compound having formula (I):(I), preferably wherein R is selected from H and COOH.

[0050] Item 4. The composition according to item 3, wherein the means for converting (3S, 11 E)- 14,16-dihydroxy-3-methyl-3,4,5,6,9, 10-hexahydro-1 H-2-benzoxacyclotetradecin-1 ,7(8H)-dione is / are an enzyme, preferably a lactonase.

[0051] Item 5. The composition according to any one of the preceding items, wherein the at least second compound is / are (E)-2,4-dihydroxy-6-(10-hydroxy-6-oxo-1-undecen-1-yl)benzoic acid and / or (E)-1-(3,5-dihydroxy-phenyl)-10-hydroxy-1-undecen-6-one.

[0052] Item 6. The composition according to any one of the preceding items, wherein the composition further comprises one or more further compounds selected from: one or more chemicals capable of modifying mycotoxin(s); one or more polypeptides capable of modifying mycotoxin(s); one or more organic absorbent(s); one or more inorganic absorbents; one or more live, inactivated, lyophilized and / or dormant microorganisms capable of modifying mycotoxin(s); one or more plant products; one or more flavoring compounds; and one or more vitamins.

[0053] Item 7. The composition according to any one of the preceding items, wherein the at least one isoflavone is selected from genistein, daidzein, glycitein and equol.

[0054] Item 8. The composition according to any one of the preceding items, wherein(i) the composition is foodstuff; fodder; feed; silage; wet distillers grain; dried distillers grain with solubles; intermediate / s thereof; and / or mixture / s thereof; and / or wherein(ii) the composition comprises cereal(s) such as rice, wheat, rye, oats, corn or maize, barley, sorghum, and / or soy or other Fabaceae plant(s) suitable for human and / or animal consumption; and / or product(s) thereof.

[0055] Item 9. Method for reducing estrogenic effect(s) and / or endocrine impact(s) of a xenoestrogen, comprising forming a mixture of the xenoestrogen with a compound having formula (I):(I), preferably wherein R is selected from H and COOH.

[0056] Item 10. The method according to item 9, wherein said forming a mixture comprises the step of contacting the xenoestrogen with the compound.

[0057] Item 11 . Method for reducing estrogenic effect(s) and / or endocrine impact(s) of a xenoestrogen on an animal cell, comprising the step of contacting the animal cell with a compound having formula (I):(I), preferably wherein R is selected from H and COOH.

[0058] Item 12. Method for manufacturing a composition (e.g. a nutritional composition), the method comprising the steps of(i) providing a compound having formula (I):(I), preferably wherein R is selected from H and COOH; and(ii) contacting the composition with the compound.

[0059] Item 13. The method according to item 12, further comprising a step (iii) of mixing the composition with the compound.

[0060] Item 14. The method according to any one of items 12-13, wherein(i) the composition is foodstuff; fodder; feed; silage; wet distillers grain; dried distillers grain with solubles; intermediate / s thereof; and / or mixture / s thereof; and / or wherein(ii) the composition comprises cereal(s) such as rice, wheat, rye, oats, corn or maize, barley, sorghum, and / or soy or other Fabaceae plant(s) suitable for human and / or animal consumption; and / or product(s) thereof.

[0061] Item 15. The method according to any one of items 12-14, wherein the composition comprises a xenoestrogen.

[0062] Item 16. The method according to any one of items 12-15, wherein the composition comprises a phytoestrogen, preferably an isoflavone, more preferably an isoflavone selected from genistein, daidzein, glycitein and equol.

[0063] Item 17. The method according to any one of items 9-16, wherein the compound is provided by providing (3S,11 E)-14,16-dihydroxy-3-methyl-3,4,5,6,9,10-hexahydro-1 H-2- benzoxacyclotetradecin-1 ,7(8H)-dione; and means for converting (3S,11 E)-14,16-dihydroxy-3- methyl-3,4,5,6,9,10-hexahydro-1 H-2-benzoxacyclotetradecin-1 ,7(8H)-dione to a compound having formula (I).

[0064] Item 18. The method according to item 17, wherein the means for converting (3S,11 E)- 14,16-dihydroxy-3-methyl-3,4,5,6,9, 10-hexahydro-1 H-2-benzoxacyclotetradecin-1 ,7(8H)-dione is / are an enzyme, preferably a lactonase.

[0065] Item 19. The method according to any one of items 9-18, wherein the compound is / are (E)-2,4-dihydroxy-6-(10-hydroxy-6-oxo-1-undecen-1-yl)benzoic acid and / or (E)-1-(3,5-dihydroxy- phenyl)-10-hydroxy-1-undecen-6-one.

[0066] Item 20. Use of a compound having formula (I):(I),preferably wherein R is selected from H and COOH; for reducing estrogenic effect(s) and / or endocrine impact(s) of a xenoestrogen.

[0067] Item 21 . Use of a compound having formula (I):(I), preferably wherein R is selected from H and COOH; for reducing estrogenic effect(s) and / or endocrine impact(s) of a composition (e.g. nutritional composition) comprising a xenoestrogen.

[0001] Item 22. The use according to any one of items 20-21 , wherein the xenoestrogen is a phytoestrogen, preferably an isoflavone, more preferably selected from genistein, daidzein, glycitein and equol.

[0068] Item 23. The use according to any one of items 20-22, wherein the compound is / are (E)- 2,4-dihydroxy-6-(10-hydroxy-6-oxo-1-undecen-1-yl)benzoic acid and / or (E)-1-(3,5-dihydroxy- phenyl)-10-hydroxy-1-undecen-6-one.

[0002] Item 24. Composition, e.g. nutritional composition, for use in treatment, amelioration and / or prevention of symptoms associated with estrogenic effect(s) and / or endocrine impact(s) of a xenoestrogen;(i) wherein the composition comprises at least a compound having formula (I):(I), preferably wherein R is selected from H and COOH; and / or ii) wherein the composition comprises (3S,11 E)-14,16-dihydroxy-3-methyl-3,4,5,6,9,10- hexahydro-1 H-2-benzoxacyclotetradecin-1 ,7(8H)-dione; and means for converting (3S,11 E)-14,16-dihydroxy-3-methyl-3,4,5,6,9, 10-hexahydro-1 H-2-benzoxacyclotetradecin-1 ,7(8H)-dione to a compound having formula (I):(I), preferably wherein R is selected from H and COOH.

[0069] Item 25. The composition for use according to item 24, wherein the means for converting (3S,11 E)-14,16-dihydroxy-3-methyl-3,4,5,6,9,10-hexahydro-1 H-2-benzoxacyclotetradecin- 1 ,7(8H)-dione is / are an enzyme, preferably a lactonase.

[0070] Item 26. The composition for use according to any one of items 24-25, wherein the compound is / are (E)-2,4-dihydroxy-6-(10-hydroxy-6-oxo-1-undecen-1-yl)benzoic acid and / or (E)- 1-(3,5-dihydroxy-phenyl)-10-hydroxy-1-undecen-6-one.

[0071] Item 27. The composition for use according to any one of items 24-26, wherein(i) the composition is foodstuff; fodder; feed; silage; wet distillers grain; dried distillers grain with solubles; intermediate / s thereof; and / or mixture / s thereof; and / or wherein(ii) the composition comprises cereal(s) such as rice, wheat, rye, oats, corn or maize, barley, sorghum, and / or soy or other Fabaceae plant(s) suitable for human and / or animal consumption; and / or product(s) thereof.

[0072] Item 28. The composition for use according to any one of items 24-27, wherein the xenoestrogen is a phytoestrogen, preferably an isoflavone, more preferably an isoflavone selected from genistein, daidzein, glycitein and equol.

[0073] In the following, the present invention is further described by non-limiting figures and examples, wherein Figure 1 shows the reduction of estrogenic effects of xenoestrogens upon addition of test substance 1 or 2 (TS1 or TS2, respectively). Estrogenic effects without added test substance (as indicated by "-") was set to 100%.

[0074] The Figures shown herein and described below serve merely as illustrative examples and are not to be construed as limiting embodiments of the present invention.Examples

[0075] The present invention as disclosed herein is not limited to specific embodiments, figures, methodology, examples, protocols etc. described herein but solely defined by the claims.Example 1

[0076] In order to assess whether a substance bears the potential and thus the risk for effecting an endocrine impact, in particular by eliciting estrogenic effect(s), Ishikawa cells were incubated with said substance. The Ishikawa cell line is a human endometrial adenocarcinoma cell line which represents a well-characterized estrogen-sensitive cell model and which produces both isoforms of the estrogen receptor, ERa and ERp. Ishikawa cells are further known to show induction of gene expression encoding alkaline phosphatase (ALP) and thus stimulation of ALP activity, upon contact with estrogenic substance(s), e.g. Miller et al. 2016. Toxical. Sci. 154(1): 162-173; Albert et al. 1990. Cancer Res. 50(11): 3306-10. Thus, the amount of ALP activity can be used to evaluate estrogenic effect(s) of test substances.

[0077] To measure estrogenicity, several substances were tested either alone or in combination, essentially as described in Grgic et al. 2022. Arch Toxicol. 96(12): 3385-3402. Stocks of Ishikawa cells were stored in liquid nitrogen containers or at -80 °C. Two weeks prior to first contact with a test substance or a xenoestrogen substance, the cells were cultivated at 37 °C, 5% CO2, 95% humidity in growth medium (minimal essential medium as described by Eagle. 1959. Science 130(3373): 432-437; supplemented with 5% v / v heat-inactivated fetal bovine serum; 1% L- glutamine; 1 % penicillin-streptomycin). Cells of passage no. 5 were split at a confluency of approximately 80%, and further cultivated to a max. passage no. of 40.

[0078] Prior to the estrogenicity assay, growth medium was replaced with assay medium (Dulbecco's Modified Eagle's Medium as described by Dulbecco et al. 1959. Science 8(3): 396- 7; supplemented with 5% charcoal-stripped fetal bovine serum; 1 % penicillin-streptomycin).

[0079] In 96-well plates, Ishikawa cells were seeded in assay medium at a concentration of 10000 cells per well. The cells were first grown for 48 h, and then incubated for additional 48 h with different concentrations of one substance or combinations of substances. Xenoestrogen substances and test substances were dissolved in DMSO and diluted in assay medium to a final concentration of 1% DMSO. A solvent control was performed by incubation with 1% DMSO in assay medium without any test substance. A positive control was performed by incubation with 1 nM of the estrogen-active hormone 17p-estradiol (E2). Experiments were performed in at least five independent biological replicates using different cell passages, with technical triplicates. After incubation, the supernatants were removed and all cells were washed three times with 150 pL ofphosphate-buffered saline (PBS) per well. For cell lysis, PBS was removed and the plates containing the washed cells were incubated at -80 °C for at least 20 min. Then, the plate was incubated at room temperature for 5 min to thaw the lysed cells. Thereafter, 50 pL of reaction buffer (5 mM 4-nitrophenylphosphate, 1 M diethanolamine, 0.24 mM MgCh, pH 9.8) was added per well, in the dark. The plate was left at room temperature for 5 min, then absorbance at 405 nm was measured for 1 h at 37 °C. ALP activity was determined as the slope of the absorbance curve, wherein the slope of the curve obtained from the solvent control was set to 0% ALP activity, and the slope of the curve obtained from the positive control was set to 100% ALP activity. A high ALP activity thus indicated high estrogenic effect.Example 2

[0080] The estrogenicity and thus endocrine impact potential of xenoestrogens was tested. In addition, test substances were studied for their potential to reduce or mitigate the estrogenicity, i.e. the estrogenic effect(s) of the xenoestrogens. As exemplary xenoestrogen substances, genistein (GEN), daidzein (DAI), glycitein (GLY) and equol (EQO) were tested at different concentrations. As test substances to reduce the estrogenic effects of these xenoestrogen substances, (E)-2,4-dihydroxy-6-(10-hydroxy-6-oxo-1-undecen-1-yl)benzoic acid was used as test substance 1 (TS1 ; compound having formula (I), wherein R is COOH); and (E)-1-(3,5- dihydroxy-phenyl)-10-hydroxy-1-undecen-6-one was used as test substance 2 (TS2; compound having formula (I), wherein R is H).

[0081] As shown in Fig. 1 , addition of as little as 0.001 nM of TS1 or TS2 reduced ALP activity, i.e. estrogenic effect, of 0.01 pM GEN by approximately 70%, of 0.1 pM GEN and of 1 pM GEN each by approximately 60%, of 0.1 pM DAI by approximately 65-85%, of 0.1 pM EQO by approximately 50-60%, and of 10 pM GLY by approximately 45-70%. Higher concentrations of 10 nM of TS1 or TS2 reduced estrogenic effect of 0.01 pM of GEN by approximately 90-100%. Similarly, 10 nM of test substances reduced estrogenic effect of 0.01 pM and 0.1 pM of EQO by approximately 90-100% and approximately 55%, respectively.

Claims

Claims1. Composition, in particular a nutritional composition, comprising at least one isoflavone, and at least a second compound having formula (I):(I), wherein R is selected from H and COOH.

2. The composition according to claim 1 , wherein the composition further comprises one or more further compounds selected from one or more chemicals capable of modifying mycotoxin(s); one or more polypeptides capable of modifying mycotoxin(s); one or more organic absorbent(s); one or more inorganic absorbents; one or more live, inactivated, lyophilized and / or dormant microorganisms capable of modifying mycotoxin(s); one or more plant products; one or more flavoring compounds; and one or more vitamins.

3. The composition according to claim 1 or 2, wherein the at least one isoflavone is selected from genistein, daidzein, glycitein and equol.

4. The composition according to any one of claims 1-3, wherein the composition is foodstuff; fodder; feed; silage; wet distillers grain; dried distillers grain with solubles; intermediate / s thereof; and / or mixture / s thereof; and / or wherein the composition comprises cereal(s) such as rice, wheat, rye, oats, corn or maize, barley, sorghum, and / or soy or other Fabaceae plant(s) suitable for human and / or animal consumption; and / or product(s) thereof.

5. Method for reducing estrogenic effect(s) of a xenoestrogen, comprising forming a mixture of the xenoestrogen with a compound having formula (I):(I), wherein R is selected from H and COOH.

6. Method for reducing estrogenic effect(s) of a xenoestrogen on an animal cell, comprising the step of contacting the animal cell with a compound having formula (I):(I), wherein R is selected from H and COOH.

7. Method for manufacturing a composition, in particular for manufacturing a nutritional composition, the method comprising the steps of providing a compound having formula (I):(I), wherein R is selected from H and COOH; and contacting the composition with the compound.

8. The method according to claim 7, wherein the composition is foodstuff; fodder; feed; silage; wet distillers grain; dried distillers grain with solubles; intermediate / s thereof; and / or mixture / s thereof;and / or wherein the composition comprises cereal(s) such as rice, wheat, rye, oats, corn or maize, barley, sorghum, and / or soy or other Fabaceae plant(s) suitable for human and / or animal consumption; and / or product(s) thereof.

9. The method according to claim 7 or 8, wherein the composition comprises a xenoestrogen.

10. The method according to claim 9, wherein the xenoestrogen is a phytoestrogen, preferably an isoflavone, more preferably an isoflavone selected from genistein, daidzein, glycitein and equol.11 . Use of a compound having formula (I):(I), wherein R is selected from H and COOH; for reducing estrogenic effect(s) of a xenoestrogen.

12. The use according to claim 11 , wherein the xenoestrogen is a phytoestrogen, preferably an isoflavone, more preferably selected from genistein, daidzein, glycitein and equol.

13. Composition, e.g. nutritional composition, for use in treatment, amelioration and / or prevention of symptoms associated with estrogenic effect(s) of a xenoestrogen; wherein the composition comprises at least a compound having formula (I):wherein R is selected from H and COOH.

14. The composition for use according to claim 13, wherein the composition is foodstuff; fodder; feed; silage; wet distillers grain; dried distillers grain with solubles; intermediate / s thereof; and / or mixture / s thereof; and / or wherein the composition comprises cereal(s) such as rice, wheat, rye, oats, corn or maize, barley, sorghum, and / or soy or other Fabaceae plant(s) suitable for human and / or animal consumption; and / or product(s) thereof.

15. The composition for use according to claim 13 or 14, wherein the xenoestrogen is a phytoestrogen, preferably an isoflavone, more preferably an isoflavone selected from genistein, daidzein, glycitein and equol.

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