WO2024223018A1

WO2024223018A1 - Bitter-masking of kaempferol or derivatives thereof

Info

Publication number: WO2024223018A1
Application number: PCT/EP2023/060580
Authority: WO
Inventors: Veronika Somoza; Katrin GRADL; Jakob Peter Ley; Silke HILLEBRAND; Maria Blings; Susanne Paetz; Rebecca WIEBUSCH
Original assignee: Symrise Ag; Leibniz Institute For Food Systems Biology At The Technical University Of Munich
Priority date: 2023-04-24
Filing date: 2023-04-24
Publication date: 2024-10-31
Also published as: WO2024223539A1

Abstract

The present invention relates to the use of a compound or a mixture of compounds for reducing or masking the bitter taste impression of kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, to a method for reducing or masking the bitter taste impression of kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, to a composition comprising such a compound or mixture of compounds and kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, and to a nutritional preparation, semi-finished preparation or pharmacological preparation comprising such a composition.

Description

Bitter-masking of Kaempferol or derivatives thereof

The present invention relates to the use of a compound or a mixture of compounds for reducing or masking the bitter taste impression of kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, to a method for reducing or masking the bitter taste impression of kaempferol or a glycoside thereof or a glycoside ester thereof or mix- tures thereof, to a composition comprising such a compound or mixture of compounds and kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, and to a nutritional preparation, semi-finished preparation or pharmacological preparation comprising such a composition.

Protein isolates are widely used in the human food industry. Particularly for providing meat- free or meat-reduced food products or components thereof, vegetable protein isolates are required in large amounts. However, also meat-including food products often contain vegetable protein isolates. Thus, such isolates are essential in the human food industry.

One common source for providing vegetable protein isolates is rapeseed (canola, Brassica napus). However, rapeseed protein isolates typically provide a disadvantageous bitter taste, which reduces the suitability of such protein isolates for being used in food products. In a recent publication, Hald et al., “Kaempferol 3-O-(2"'-O-Sinapoyl-[3-sophoroside) Causes the Undesired Bitter Taste of Canola/Rapeseed Protein Isolates", J Agric Food Chem, 2019 Jan 9;67(1):372-378 discovered that Kaempferol and its glycosides or glycoside esters, particularly the glucoside kaempferol 3-O-sinapoyl-sophoroside 7-O-glucoside (K3OSS), are responsible for providing the bitter taste in rapeseed protein isolates.

Kaempferol 3- O-(2"'- O-Sinapoyl-fi-sophoroside)

Kaempferol (3,4',5,7-tetrahydroxyflavone) is a natural flavonoid, which occurs in several vegetables and fruits such as apples, grapes, tomatoes, green tea, potatoes, onions, broc- coli, Brussels sprouts, squash, cucumbers, lettuce, green beans, peaches, blackberries, raspberries, and spinach.

Kaempferol Thus, the presence of kaempferol or a glycoside thereof or a glycoside ester in such protein isolates may provide a bitter taste, which should be reduced or avoided for using such protein isolates in food products.

There is thus a need for options to reduce or avoid the bitter taste of such protein isolates, particularly rapeseed protein isolates.

The primary object of the present invention was thus to provide possibilities for reducing or masking the bittertaste impression of kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof.

The primary object of the present invention is solved by the use of a compound or a mixture of compounds wherein the compound or, respectively, one or two or more or all compounds in the mixture of compounds is/are selected from the group consisting of eriodictyol, hesperetin, naringenin and phloretin, for reducing or masking the bitter taste impression of kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof.

It was surprisingly found that eriodictyol, hesperetin, naringenin and phloretin provided a particularly strong reduction or masking effect with regard to the bitter taste impression of kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof.

Several substances are known as bitter-masking substances. Such frequently used substances include eriodictyol, homoeriodictyol, naringenin, hesperetin, matairesinol and phloretin.

For example, eriodictyol has been described for masking the bitter taste of several different bitter tasting substances, such as in EP 1 258 200, EP 1 972 203, WO 2017/088936, WO 2017/182538 or WO 2020/035177. However, a bitter-masking effect with regard to kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof has not been described.

Likewise, homoeriodictyol, naringenin, hesperetin, matairesinol and phloretin have also been described for masking the bitter taste of several different bitter tasting substances. However, a bitter-masking effect with regard to kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof has not been described.

It was surprisingly found that only some of these substances are able to reduce or mask the bitter taste impression of kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof.

Furthermore, it was surprisingly found that eriodictyol provided the strongest reduction or bitter-masking effect compared to all tested substances. This was particularly surprising, since the structurally highly similar substance homoeriodictyol did not provide such an effect. In contrast, homoeriodictyol provided a very low or even no reduction or bitter-masking effect.

Eriodictyol Homoeriodictyol

The term “kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof’, as used herein, refers to kaempferol or a glycoside of kaempferol or a glycoside ester of kaempferol or to mixtures including one or two or all of the compounds selected from the group consisting of kaempferol, glycosides of kaempferol and glycoside esters of kaempferol.

The term “a glycoside”, as used herein, is to be understood as one or more glycoside(s). The same applies accordingly to the term “a glycoside ester”.

Preferred examples for kaempferol glycosides or glycoside esters are shown in the following formulas:

Kaempferol 3-sophoroside-7-0-glucoside

7-(p-D-Glucopyranosyloxy)-5-hydroxy-3-[[2-0-[2-0-[(2E)-3-(4-hydroxy-3,5-dimethoxy- phenyl)-1-oxo-2-propen-1-yl]-p-D-glucopyranosyl]-p-D-glucopyranosyl]oxy]-2-(4-hydroxy- phenyl)-4H-1 -benzopyran-4-one

(Kaempferol 3-0-(2'"-0-sinapoyl-p-sophoroside)-7-0-glucoside)

5,7-Dihydroxy-2-(4-hydroxyphenyl)-3-[[2-0-[2-0-(3,4,5-trihydroxybenzoyl)-p-D-glucopyra- nosyl]-p-D-glucopyranosyl]oxy]-4/7-1-benzopyran-4-one

(Kaempferol 3-0-(2"'-0-galloyl-p-sophoroside))

,7-Dihydroxy-2-(4-hydroxyphenyl)-3-[[2-0-[2-0-[(2E)-3-(4-hydroxyphenyl)-1-oxo-2-pro- pen-1-yl]-p-D-glucopyranosyl]-p-D-glucopyranosyl]oxy]-4H-1-benzopyran-4-one

(Kaempferol 3-0-(2"'-0-coumaroyl-p-sophoroside))

-[[2-0-[6-0-[(2E)-3-(3,4-Dihydroxyphenyl)-1-oxo-2-propen-1-yl]-p-D-glucopyranosyl]-p-

D-glucopyranosyl]oxy]-5,7-dihydroxy-2-(4-hydroxyphenyl)-4H-1-benzopyran-4-one

(Kaempferol 3-0-(2"'-0-caffeoyl-p-sophoroside))

5,7-Dihydroxy-3-[[2-0-[2-0-[(2E)-3-(4-hydroxy-3-methoxyphenyl)-1-oxo-2-propen-1-yl]-p-

D-glucopyranosyl]-p-D-glucopyranosyl]oxy]-2-(4-hydroxyphenyl)-4H-1-benzopyran-4-one

(Kaempferol 3-0-(2"'-0-feruloyl-p-sophoroside)). Preferably, the, one, two, three or more or all kaempferol glycosides and glycoside esters is/are selected from the group consisting of Kaempferol 3-O-sophoroside, Kaempferol 3- sophoroside-7-O-glucoside, kaempferol 3-0-(2"'-0-sinapoyl-p-sophoroside), kaempferol 3- O-sinapoyl-sophoroside 7-O-glucoside (K3OSS), Kaempferol 3-0-(2"'-0-sinapoyl-p- sophoroside)-7-0-glucoside, Kaempferol 3-0-(2"'-0-galloyl-p-sophoroside), Kaempferol 3- 0-(2"'-0-coumaroyl-p-sophoroside)), Kaempferol 3-0-(2"'-0-caffeoyl-p-sophoroside), Kaempferol 3-0-(2"'-0-feruloyl-p-sophoroside).

Preferably, the, one, two, three or more or all kaempferol glycosides and glycoside esters is/are selected from kaempferol glucosides and glucoside esters. Particularly preferably, the, one, two or three or all kaempferol glucosides and glucoside esters is/are selected from the group consisting of kaempferol 3-O-sinapoyl-sophoroside 7-O-glucoside (K3OSS), kaempferol 3-O-sophoroside, and kaempferol 3-0-(2"'-0-sinapoyl-p-sophoro- side)-7-0-glucoside.

Especially preferably, the kaempferol glycoside ester, as described herein, is K3OSS. It is thus preferred that the use according to the invention is for reducing or masking the bitter taste impression of one or more kaempferol glycoside(s) or glycoside ester(s) or a mixture comprising one or more kaempferol glycoside(s) and/or glycoside ester(s), wherein the, one, two, or three or all kaempferol glycosides or glycoside ester(s) is/are selected from the group consisting of Kaempferol 3-O-sophoroside, Kaempferol 3-sophoroside-7-0- glucoside, kaempferol 3-0-(2"'-0-sinapoyl-p-sophoroside), kaempferol 3-O-sinapoyl- sophoroside 7-O-glucoside (K3OSS), Kaempferol 3-0-(2"'-0-sinapoyl-p-sophoroside)-7-0- glucoside, Kaempferol 3-0-(2"'-0-galloyl-p-sophoroside), Kaempferol 3-0-(2"'-0-couma- royl-p-sophoroside)), Kaempferol 3-0-(2"'-0-caffeoyl-p-sophoroside), Kaempferol 3-O-(2"'- O-feruloyl-p-sophoroside.

It is further preferred that the use according to the invention is for reducing or masking the bitter taste impression of one or more kaempferol glycoside(s) or glycoside ester(s) or a mixture comprising one or more kaempferol glycoside(s) and/or glycoside ester(s), wherein the, one, two, or all kaempferol glycoside or glycoside ester is selected from the group consisting of kaempferol glucosides or glucoside esters, particularly preferably selected from the group consisting of kaempferol 3-O-sinapoyl-sophoroside 7-O-glucoside (K3OSS), kaempferol 3-O-sophoroside, and kaempferol 3-0-(2"'-0-sinapoyl-p-sophoro- side)-7-0-glucoside.

It is further preferred that the use according to the invention is for reducing or masking the bitter taste impression of one or more kaempferol glycoside(s) or a mixture comprising one or more kaempferol glycoside(s), preferably wherein the, one, two, three or more or all kaempferol glycoside(s) is/are selected from the kaempferol glycosides listed above, preferably from the kaempferol glucosides listed above.

It is further preferred that the use according to the invention is for reducing or masking the bitter taste impression of one or more kaempferol glycoside ester(s) or a mixture comprising one or more kaempferol glycoside ester(s), preferably wherein the, one, two, three or more or all kaempferol glycoside ester(s) is/are selected from the kaempferol glycoside esters listed above, preferably from the kaempferol glucoside esters listed above.

As described above, it was surprisingly found that only some of the tested substances, which are typically used for masking a bitter taste of other substances, are able to reduce or mask the bitter taste impression of kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof. It is thus preferred in the use according to the invention that the compound or, respectively, one compound in the mixture is eriodictyol.

Preferably, the term “eriodictyol”, as used herein, refers to the R-eriodictyol enantiomer, the S-eriodictyol enantiomer or a mixture thereof. Preferably, the term “eriodictyol”, as used herein, refers to a mixture of the R- and S-enantiomers, preferably wherein the mixture comprises 0.01 to 99.99 wt.-%, preferably 0.1 to 99.9 wt.-%, preferably 1 to 99 wt.-%, preferably 5 to 95 wt.-%, preferably 10 to 90 wt.-%, preferably 20 to 80 wt.-%, preferably 25 to 75 wt.-%, preferably 30 to 70 wt.-%, preferably 40 to 60 wt.-%, R-eriodictyol, based on the summed weight of R- and S-eriodictyol in the mixture. Preferably, the term “eriodictyol”, as used herein, refers to a mixture of the R- and S-enantiomers, preferably wherein the mixture comprises 0.01 to 99.99 wt.-%, preferably 0.1 to 99.9 wt.-%, preferably 1 to 99 wt.-%, preferably 5 to 95 wt.-%, preferably 10 to 90 wt.-%, preferably 20 to 80 wt.-%, preferably 25 to 75 wt.-%, preferably 30 to 70 wt.-%, preferably 40 to 60 wt.-%, S-eriodictyol, based on the summed weight of R- and S-eriodictyol in the mixture.

It is thus preferred in the use according to the invention that the compound or, respectively, one compound in the mixture is hesperetin.

Preferably, the term “hesperetin”, as used herein, refers to the R-hesperetin enantiomer, the S- hesperetin enantiomer or a mixture thereof. Preferably, the term “hesperetin”, as used herein, refers to a mixture of the R- and S-enantiomers, preferably wherein the mixture comprises 0.01 to 99.99 wt.-%, preferably 0.1 to 99.9 wt.-%, preferably 1 to 99 wt.-%, preferably 5 to 95 wt.-%, preferably 10 to 90 wt.-%, preferably 20 to 80 wt.-%, preferably 25 to 75 wt.-%, preferably 30 to 70 wt.-%, preferably 40 to 60 wt.-%, R- hesperetin, based on the summed weight of R- and S- hesperetin in the mixture. Preferably, the term “hesperetin”, as used herein, refers to a mixture of the R- and S-enantiomers, preferably wherein the mixture comprises 0.01 to 99.99 wt.-%, preferably 0.1 to 99.9 wt.-%, preferably 1 to 99 wt.-%, preferably 5 to 95 wt.-%, preferably 10 to 90 wt.-%, preferably 20 to 80 wt.- %, preferably 25 to 75 wt.-%, preferably 30 to 70 wt.-%, preferably 40 to 60 wt.-%, S- hesperetin, based on the summed weight of R- and S- hesperetin in the mixture.

It is thus preferred in the use according to the invention that the compound or, respectively, one compound in the mixture is naringenin.

Preferably, the term “naringenin”, as used herein, refers to the R-naringenin enantiomer, the S-naringenin enantiomer or a mixture thereof. Preferably, the term “naringenin”, as used herein, refers to a mixture of the R- and S-enantiomers, preferably wherein the mixture comprises 0.01 to 99.99 wt.-%, preferably 0.1 to 99.9 wt.-%, preferably 1 to 99 wt.-%, preferably 5 to 95 wt.-%, preferably 10 to 90 wt.-%, preferably 20 to 80 wt.-%, preferably 25 to 75 wt.-%, preferably 30 to 70 wt.-%, preferably 40 to 60 wt.-%, R-naringenin, based on the summed weight of R- and S-naringenin in the mixture. Preferably, the term “naringenin”, as used herein, refers to a mixture of the R- and S-enantiomers, preferably wherein the mixture comprises 0.01 to 99.99 wt.-%, preferably 0.1 to 99.9 wt.-%, preferably 1 to 99 wt.-%, preferably 5 to 95 wt.-%, preferably 10 to 90 wt.-%, preferably 20 to 80 wt.- %, preferably 25 to 75 wt.-%, preferably 30 to 70 wt.-%, preferably 40 to 60 wt.-%, S- naringenin, based on the summed weight of R- and S-naringenin in the mixture.

It is thus preferred in the use according to the invention that the compound or, respectively, one compound in the mixture is phloretin.

Furthermore, it is preferred in the use according to the invention that the mass ratio of i) the compound(s), selected from the group consisting of eriodictyol, hesperetin, naringenin, phloretin and mixtures thereof, and ii) kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof is in a range of from 1 :5 to 20:1 , preferably in the range of from 1 :2.5 to 15:1 , preferably in the range of from 1 :1 to 10:1 , preferably in the range of from 2.5:1 to 7.5 to 1.

Preferably, the term “the mass ratio of eriodictyol and kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof’, as used herein, is to be understood such that for determining the mass ratio, only the mass of eriodictyol but not the mass of hesperetin, naringenin or phloretin is considered. The same applies accordingly to hesperetin, naringenin and phloretin.

Preferably, the term “the mass ratio of the compound(s), selected from the group consisting of eriodictyol, hesperetin, naringenin, phloretin and mixtures thereof, and kaempferol”, as used herein, is to be understood such that for determining the mass ratio, only the mass of kaempferol but not the mass of a kaempferol glycoside or a kaempferol glycoside ester is considered. The same applies accordingly to the kaempferol glycosides and the kaempferol glycoside esters, wherein in case it is referred to kaempferol glycosides or, respectively, kaempferol glycoside esters, the summed mass of all present kaempferol glycosides or, respectively, kaempferol glycoside esters is considered for determining the mass ratio, and wherein in case it is referred to a specific kaempferol glycoside or, respectively, kaempferol glycoside ester, only the mass of said glycoside or glycoside ester is considered. Preferably, the term “the mass ratio of a mixture of eriodictyol, hesperetin, naringenin and phloretin, and kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof’, as used herein, is to be understood such that for determining the mass ratio, the summed mass of eriodictyol, hesperetin, naringenin and phloretin is considered. In case one or two or three of eriodictyol, hesperetin, naringenin and phloretin is not present in said mixture, the mass of the respective compound(s) is considered 0 g.

Preferably, the term “the mass ratio of the compound(s), selected from the group consisting of eriodictyol, hesperetin, naringenin, phloretin and mixtures thereof, and a mixture of kaempferol or a glycoside thereof or a glycoside ester thereof’, as used herein, is to be understood such that for determining the mass ratio, the summed mass of kaempferol, a glycoside thereof and a glycoside ester thereof is considered. In case one or two of kaempferol, a glycoside thereof and a glycoside ester thereof is not present in said mixture, the mass of the respective compound(s) is considered 0 g.

Furthermore, the present invention relates to a method for reducing or masking the bitter taste impression of kaempferol or a glycoside thereof or a mixture thereof, comprising the following steps i) providing kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, ii) providing eriodictyol, hesperetin, naringenin, phloretin, or a mixture thereof, iii) mixing the compound or mixture provided in step i) with the compound or mixture provided in step ii).

What was said above with regard to kaempferol, the kaempferol glycoside, the kaempferol glycoside ester, the mixtures thereof, eriodictyol, hesperetin, naringenin, phloretin and the mixtures thereof, applies accordingly for the method according to the invention.

Preferably, providing kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof in step i) refers to the kaempferol or glycoside thereof or glycoside ester thereof or mixtures thereof, of which a bitter taste shall be reduced or masked.

Preferably, the method according to the invention further comprises the steps i.a) measuring a bitter taste impression of the kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, as provided in step i), and iv) measuring the bitter taste impression of the mixture obtained after step iii), and v) comparing the measurements obtained in steps i.a) and iv).

Preferably, in case a mixture comprising kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, is provided in step i) of the method according to the invention, the measurement in step i.a) refers to the bitter taste impression of said mixture comprising kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof.

It is preferred that the method according to the invention is for reducing or masking the bitter taste impression of one or more kaempferol glycoside(s) or glycoside ester(s) or a mixture comprising one or more kaempferol glycoside(s) and/or glycoside ester(s), wherein the, one, two, or three or all kaempferol glycosides or glycoside ester(s) is/are selected from the group consisting of Kaempferol 3-O-sophoroside, Kaempferol 3-sophoroside-7-0- glucoside, kaempferol 3-0-(2"'-0-sinapoyl-p-sophoroside), kaempferol 3-O-sinapoyl- sophoroside 7-O-glucoside (K3OSS), Kaempferol 3-0-(2"'-0-sinapoyl-p-sophoroside)-7-0- glucoside, Kaempferol 3-0-(2"'-0-galloyl-p-sophoroside), Kaempferol 3-0-(2"'-0-couma- royl-p-sophoroside)), Kaempferol 3-0-(2"'-0-caffeoyl-p-sophoroside), Kaempferol 3-O-(2"'- O-feruloyl-p-sophoroside.

It is further preferred that the method according to the invention is for reducing or masking the bitter taste impression of one or more kaempferol glycoside(s) or glycoside ester(s) or a mixture comprising one or more kaempferol glycoside(s) and/or glycoside ester(s), wherein the, one, two, or all kaempferol glycoside or glycoside ester is selected from the group consisting of kaempferol glucosides or glucoside esters, particularly preferably selected from the group consisting of kaempferol 3-O-sinapoyl-sophoroside 7-O-glucoside (K3OSS), kaempferol 3-O-sophoroside, and kaempferol 3-0-(2"'-0-sinapoyl-p-sophoro- side)-7-0-glucoside. It is further preferred that the method according to the invention is for reducing or masking the bitter taste impression of one or more kaempferol glycoside(s) or a mixture comprising one or more kaempferol glycoside(s), preferably wherein the, one, two, three or more or all kaempferol glycoside(s) is/are selected from the kaempferol glycosides listed above, preferably from the kaempferol glucosides listed above.

It is further preferred that the method according to the invention is for reducing or masking the bitter taste impression of one or more kaempferol glycoside ester(s) or a mixture comprising one or more kaempferol glycoside ester(s), preferably wherein the, one, two, three or more or all kaempferol glycoside ester(s) is/are selected from the kaempferol glycoside esters listed above, preferably from the kaempferol glucoside esters listed above.

Furthermore, it is preferred that in step ii) eriodictyol or a mixture comprising eriodictyol is provided.

Furthermore, it is preferred that in step ii) hesperetin or a mixture comprising hesperetin is provided.

Preferably, the term “hesperetin”, as used herein, refers to the R-hesperetin enantiomer, the S- hesperetin enantiomer or a mixture thereof. Preferably, the term “hesperetin”, as used herein, refers to a mixture of the R- and S-enantiomers, preferably wherein the mixture comprises 0.01 to 99.99 wt.-%, preferably 0.1 to 99.9 wt.-%, preferably 1 to 99 wt.-%, preferably 5 to 95 wt.-%, preferably 10 to 90 wt.-%, preferably 20 to 80 wt.-%, preferably 25 to 75 wt.-%, preferably 30 to 70 wt.-%, preferably 40 to 60 wt.-%, R- hesperetin, based on the summed weight of R- and S- hesperetin in the mixture. Preferably, the term “hes- peretin”, as used herein, refers to a mixture of the R- and S-enantiomers, preferably wherein the mixture comprises 0.01 to 99.99 wt.-%, preferably 0.1 to 99.9 wt.-%, preferably 1 to 99 wt.-%, preferably 5 to 95 wt.-%, preferably 10 to 90 wt.-%, preferably 20 to 80 wt.- %, preferably 25 to 75 wt.-%, preferably 30 to 70 wt.-%, preferably 40 to 60 wt.-%, S- hesperetin, based on the summed weight of R- and S- hesperetin in the mixture.

Furthermore, it is preferred that in step ii) naringenin or a mixture comprising naringenin is provided.

Furthermore, it is preferred that in step ii) phloretin or a mixture comprising phloretin is provided.

It is further preferred in the method according to the invention that the mass ratio of eriodictyol, hesperetin, naringenin, phloretin or a mixture thereof, as provided in step ii), and kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, as provided in step i), is in a range of from 1 :5 to 20:1 , preferably in the range of from 1 :2.5 to 15:1 , preferably in the range of from 1 :1 to 10:1 , preferably in the range of from 2.5:1 to 7.5 to 1.

What was said above with regard to the mass ratio and its determination, applies accordingly.

The present invention further relates to a composition comprising i) eriodictyol, hesperetin, naringenin, phloretin, or a mixture thereof, and ii) kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, preferably a protein isolate comprising kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, wherein the amount of eriodictyol, hesperetin, naringenin, phloretin, or a mixture thereof is sufficient to reduce or mask the bitter taste impression of kaempferol or a glycoside thereof or a mixture thereof.

It is preferred in the composition according to the invention that the mass ratio of i) eriodictyol, hesperetin, naringenin, phloretin or a mixture thereof, and ii) kaempferol or a glycoside thereof or a glycoside ester thereof or a mixture thereof in the composition is in a range of from 1 :5 to 20:1 , preferably in the range of from 1 :2.5 to 15:1 , preferably in the range of from 1 :1 to 10:1 , preferably in the range of from 2.5:1 to 7.5 to 1 .

It is preferred in the composition according to the invention that the amount of component ii) in the composition according to the invention is sufficient for imparting or enhancing a bitter taste impression of the composition.

It was found in Hald et al. 2019 (as described above) that a concentration of 3.4 pmol/L (corresponding to 0,000277 wt.-%) of K3OSS, is sufficient to impart a bitter taste impression.

It is thus preferred that the amount of component ii) in the composition according to the invention is at least 0.0003 wt.-%, preferably at least 0.0004 wt.-%, preferably at least 0.0005 wt.-%, preferably at least 0.001 wt.-%, preferably at least 0.005 wt.-%, preferably at least 0.01 wt.-%, preferably at least 0.05 wt.-%, preferably at least 0.1 wt.-%, preferably at least 0.5 wt.-%, based on the total weight of the composition.

Likewise, it is preferred in the use according to the invention that kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, of which the bitter taste impression shall be reduced or masked, is present in a composition comprising or consisting of said kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, wherein the composition comprises at least 0.0003 wt.-%, preferably at least 0.0004 wt.-%, preferably at least 0.0005 wt.-%, preferably at least 0.001 wt.-%, preferably at least 0.005 wt.-%, preferably at least 0.01 wt.-%, preferably at least 0.05 wt.-%, preferably at least 0.1 wt.-%, preferably at least 0.5 wt.-%, of the kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, based on the total weight of the composition.

Likewise, it is preferred in the method according to the invention that kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, provided in step i) of the method according to the invention, is provided as a composition comprising or consisting of said kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, wherein the composition comprises at least 0.0003 wt.-%, preferably at least 0.0004 wt.- %, preferably at least 0.0005 wt.-%, preferably at least 0.001 wt.-%, preferably at least 0.005 wt.-%, preferably at least 0.01 wt.-%, preferably at least 0.05 wt.-%, preferably at least 0.1 wt.-%, preferably at least 0.5 wt.-%, of the kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, based on the total weight of the composition.

In case said composition, described above in connection with the use or method according to the invention, or in case the composition according to the invention comprises more than one compound selected from kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, e.g. two or more glycosides of kaempferol or a glycoside of kaempferol and a glycoside ester of kaempferol, it is preferred that for determining the wt.-% all compounds selected from kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, which are present in said composition, are considered.

What was said above with regard to kaempferol, the kaempferol glycoside, the kaempferol glycoside ester, the mixtures thereof, eriodictyol, hesperetin, naringenin, phloretin and the mixtures thereof, applies accordingly for the composition according to the invention.

Preferably, component ii) of the composition according to the invention is or comprises K3OSS.

Preferably, component ii) of the composition according to the invention is or comprises a protein isolate comprising or consisting of K3OSS.

Preferably, the term “protein isolate”, as used herein, is a vegetable protein isolate, particularly preferably a protein isolate obtained from a fruit or vegetable selected from the group consisting of apples, blackberries, broccoli, Brussels sprouts, cucumbers, grapes, green beans, green tea, lettuce, onions, peaches, potatoes, rapeseed, raspberries, spinach, squash, tomatoes, and mixtures thereof, particularly preferably a rapeseed protein isolate. Preferably, the term “the amount of eriodictyol, hesperetin, naringenin, phloretin, or a mixture thereof is sufficient”, as used herein, refers to the mass ratio of mass ratio of i) the compound(s), selected from the group consisting of eriodictyol, hesperetin, naringenin, phloretin and mixtures thereof, and ii) kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof.

Preferably, the term “wherein the amount of eriodictyol, hesperetin, naringenin, phloretin, or a mixture thereof is sufficient to reduce or mask the bitter taste impression of kaempferol or a glycoside thereof or a mixture thereof’ refers to an amount, which is sufficient for a reduction and/or masking of at least 5 %, preferably at least 10 %, preferably at least 15 %, preferably at least 20 %, preferably at least 25 %, preferably at least 30 % of the bitter taste impression. Typically, the reduction and/or masking and its extent is determined by tasting with panellists, which is well-known to a skilled person.

Furthermore, the present invention relates to a nutritional preparation, a semi-finished preparation or a pharmacological preparation comprising a composition according to the invention.

Preferably, the amount of eriodictyol, hesperetin, naringenin, phloretin or a mixture thereof in such a preparation is at least 0.0001 wt.-%, preferably at least 0.001 wt.-%, preferably at least 0.0025 wt.-%, based on the total weight of the preparation.

Preferably, the amount of eriodictyol, hesperetin, naringenin, phloretin or a mixture thereof in such a preparation is in the range of from 0.0001 to 0.025 wt.-%, preferably in the range of from 0.001 to 0.015 wt.-%, preferably in the range of from 0.0025 to 0.01 wt.-%, based on the total weight of the preparation.

Preferably, the amount of kaempferol or a glycoside thereof or a glycoside ester thereof or a mixture thereof in such a preparation is at least 0.0001 wt.-%, preferably at least 0.003 wt.-%, preferably at least 0.0025 wt.-%, based on the total weight of the preparation. Preferably, the amount of kaempferol or a glycoside thereof or a glycoside ester thereof or a mixture thereof in such a preparation is in the range of from 0.0003 to 0.025 wt.-%, preferably in the range of from 0.001 to 0.015 wt.-%, preferably in the range of from 0.0025 to 0.01 wt.-%, based on the total weight of the preparation.

What was said above with regard to the composition according to the invention, applies accordingly to the preparation according to the invention.

What was said above with regard to the mass ratios and its determination, applies accordingly. For example, it is preferred that mass ratios described as preferred for the composition according to the invention also apply and are also preferred for the preparation according to the invention.

Preferably, nutritional preparations according to the invention are selected from baked goods (e.g. bread, dry biscuits, cakes, other pastries), confectionery (for example chocolates, chocolate bar products, other bar products, fruit gums, hard and soft caramels, chewing gum), alcoholic or non-alcoholic beverages, alcoholic beverages (e.g. cocoa, coffee, green tea, black tea, (green, black) tea extracts enriched with green, black tea, rooibos tea, other herbal teas, wine, wine-based beverages, beer, beer-containing beverages, liqueurs, schnapps, brandies, fruit-based sodas, isotonic drinks, soft drinks, nectars, fruit and vegetable juices, fruit or vegetable juice preparations), instant drinks (e.g. instant cocoa drinks, instant tea drinks, instant coffee drinks), meat products (e.g. ham, fresh sausage or raw sausage preparations, spiced or marinated fresh or cured meat products), eggs or egg products (dry egg, egg white, egg yolk), cereal products (e.g. breakfast cereals, muesli bars, pre-cooked finished rice products), dairy products (e.g. full-fat or reduced-fat milk drinks, rice pudding, yoghurt, kefir, cream cheese, soft cheese, hard cheese, dried milk powder, whey, butter, buttermilk, partially or completely hydrolysed milk protein containing products), products of soy protein or other soybean fractions (e.g. soymilk and products made therefrom, beverages containing soy protein or enzymatically treated, drinks containing soybean meal, soybean lecithin-containing preparations, fermented products such as tofu or tempe or products made therefrom and mixtures with fruit preparations and optional flavors), fruit preparations (e.g. jams, fruit ice creams, fruit sauces, fruit fillings), vegetable preparations (e.g. ketchup, sauces, dried vegetables, frozen vegetables, pre-cooked vegetables, cooked vegetables), snacks (e.g. baked or fried potato chips or car potato dough products, corn- or peanut-based extrudates), products based on fat and oil or emulsions thereof (e.g. mayonnaise, remoulade, dressings, in each case full-fat or reduced-fat), other prepared meals and soups (e.g. dry soups, instant soups, pre-cooked soups), spices, condiments and in particular seasonings, which are used, for example, in the snack area, sweetener preparations, tablets or sachets, other preparations for sweetening or whitening beverages or other foodstuffs.

Preferably, a semi-finished product refers to a substance, product or good that has not completed the manufacturing or production process and is not ready for sale to or use or consumption by consumers and/or clients in the food service or catering industry. Thus, a semi-finished product preferably refers to a product, good or article that is intended for further industrial processing and/or use in industrial processing.

Preferably, a semi-finished product is to be understood as a product which is unsuitable for use as a ready-to-eat foodstuff. Only by mixing with at least one other ingredient (i.e., by reducing the concentration of the flavouring composition) and optionally further processing steps (e.g., heating, freezing) is the semi-finished product converted into a ready-to-eat food.

Preferably, semi-finished products are selected from substances, products or goods intended for use in food products, such as extracts, mash, powder, oil or paste made from vegetables or fruit, spices, toppings, seasoning mixes, granulated or pulverized foodstuffs.

Preferably, pharmacological preparations according to the invention are in the form of capsules, tablets (uncoated and coated tablets, for example having coatings resistant to gastric juices), sugar-coated tablets, granules, pellets, mixtures of solids, dispersions in liquid phases, as emulsions, as powders, as solutions, as pastes or as other swallowable or chewable preparations and are preferably used as medicines only available by prescription, from pharmacies or other medicines or as food supplements.

Fig. 1 shows the results of the proton secretion assay of Example 2 with eriodictyol (A), homoeriodictyol (HED, B), phloretin (C), naringenin (D), matairesinol (E) and hesperetin (F), data are presented as mean IPX ± SEM differences from K3OSS only treated cells (IPX = 0), with *: p < 0.0332; **: p < 0.0021 ; ***: p < 0.0002; and ****: p < 0.0001 via Ordinary One Way ANOVA test using a Dunnett’s Multiple Comparisons Test.

Further aspects and advantages of the invention result from the subsequent description of preferred examples. Examples

Example 1 : Kaempferol and its glycosides or glycoside esters as bitter substance

According to the publication “Characterization of Bitter Compounds via Modulation of Proton Secretion in Human Gastric Parietal Cells in Culture” by K. I. Liszt, J. Hans, J. P. Ley, E. Kock and V. Somoza J. Agric Food Chem 2018 Vol. 66 Issue 10 Pages 2295-2300, the cell model HGT-1 is used as a prediction tool for bitter taste and bitter taste agonists.

Therefore in Example 1 the activity of K3OSS on the HGT-1 model and in Example 2 the combined activities of K3OSS with potential antagonists are shown.

A total of 1 ,000,000 viable HGT-1 cells were spread in a 6-well plate and allowed to settle for 24 h at 37°C, 95% humidity, and 5% CO2. After incubation with 6.8 pm K3OSS for 30 min, RNA was isolated using the pegGOLD RNA Kit. Quantity and guality of RNA were checked spectrophotometrically at a wavelength of 260 nm and by calculation of the absorbance ratio at 260 and 280 nm wavelength using a NanoDrop One (Thermo Fisher Scientific Inc., USA). Removal of gDNA and synthesis of cDNA were performed using the iScript gDNA Clear cDNA Synthesis Kit following the manufacturer’s protocol. Real-time- gPCR (RT-gPCR) was performed with 50 ng cDNA amplified with SsoAdvanced Universal SYBR Green Supermix. Peptidylprolyl isomerase A (PPIA) and glyceraldehyde 3-phos- phate dehydrogenase (GAPDH) were used as reference genes.

RT-gPCR analyses of TAS2Rs gene expression after exposure of HGT-1 cells to 6.8 pM of K3OSS resulted in an up-regulation of TAS2Rs as compared to treatment of the cells with Krebs-Ringer-Hepes-Buffer (KRHB, control). mRNA levels of the bitter receptors TAS2R3, 4, 5, 13, 8, 13, 30, 31 , 39, 40, 43, 45, 46, and 50 were higher in K3OSS treated cells than in controls. Thus, K3OSS was confirmed as bitter tasting substance.

Example 2: In-vitro bitter masking of K3OSS by candidates

The intracellular pH, calculated as intracellular proton index (IPX) as indicator of cellular proton secretion linked to bitter taste receptor (TAS2R) regulation, was measured in HGT- 1 cells by means of the pH-sensitive fluorescence dye 1 ,5 carboxy-seminaphto-rhodafiuor acetoxymethylester (SNARF-1-AM), as described in Liszt et al, “Characterization of Bitter Compounds via Modulation of Proton Secretion in Human Gastric Parietal Cells in Culture", J. Agric. Food Chern. 2018, 66, 2295-2300. 100,000 HGT-1 cells were seeded in a black 96-well plate. After 24 h, cells were stained with 3 pM SNARF-1-AM for 30 min at standard cell culture conditions and co-incubated for 10 min with 17 pM K3OSS and the different candidates (homoeriodictyol, eriodicytol, hes- peretin, naringenin, phloretin, matairesinol) in different concentrations.

Histamine (1 mM) was used as internal reference, whereas HGT-1 cells exposed to Krebs- Ringer-Hepes-Buffer (KRHB) only were used as control. Fluorescence was measured at 580 and 640 nm emission after excitation at 488 nm, by means of a Flexstation 3 (Molecular Devices, California, USA). Using a nigericin calibration curve, the intracellular pH and the resulting intracellular H⁺ concentration were calculated. Hence, the ratio between treated and nontreated cells (KRHB only) was calculated and Iog2 transformed to determine the intracellular proton index (IPX).

The impact of the co-treatments of K3OSS and the above candidates on TAS2R-linked proton secretion by HGT-1 cells was analyzed by quantitation of the intracellular proton index (IPX). A negative IPX value indicates an increased proton secretion and potential activation of TAS2Rs (indicating a bitter taste), a positive IPX value indicates a reduced proton secretion and potential de-activation of TAS2Rs (indicating a reduced and/or masked bitter taste).

A positive IPX value was obtained for co-treatment with eriodictyol, naringenin or phloretin, already at low concentrations. Further, a positive IPX value was obtained for co-treatment with hesperetin. Surprisingly, no positive IPX value was obtained for co-treatment with homoeriodictyol, although the structurally similar candidate eriodictyol provided a positive IPX value. Similarly, no positive IPX value was obtained for co-treatment with matairesinol.

Thus, eriodictyol, naringenin and phloretin provided a particularly positive IPX and hesperetin provided a slightly positive IPX, indicating a reduction and/or masking of the bitter taste impression of K3OSS. The results are depicted in Fig. 1.

Example 3: Sensory analysis of the candidates’ bitter taste reduction and/or masking

A base including 20 mg/kg K3OSS was prepared. Samples including the base only or the base in addition to the different candidates were prepared. Concentrations of candidates were selected to avoid disturbing side tastes. The samples were analysed by a trained panel (n=20) with regard to their bitter taste. During the tests (bitter-duo tests), the panellists wear nose-clips to exclude any olfactory impressions. The bitter taste was evaluated by the panellists. Subsequently, the evaluation of the samples including a candidate was compared with the evaluation of the base without a candidate, to obtain the reduction or, respectively, masking of the bitter taste impression

Example 4: Protein extrudate

Extrudates were prepared by dry mixing of the solid ingredients, addition of water (factor 12:8 water to solid by weight) and kneeding and heating in a commercial twin-screw extruder with an amount of 15 kg/h at heating temperature of 170°C (mass temperature about 128-130°C). The extrudate is cooled down on the extruder nozzle (45 °C). The cooled extrudates were cut to chunks and backed together with 7 wt. % sunflower oil at 175°C air heat and 30% humidity for 8 min.

Claims

1 . Use of a compound or a mixture of compounds wherein the compound or, respectively, one or two or more or all compounds in the mixture of compounds is/are selected from the group consisting of eriodictyol, hes- peretin, naringenin and phloretin, for reducing or masking the bitter taste impression of kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof.

2. Use according to claim 1 , for reducing or masking the bitter taste impression of one or more kaempferol glycoside(s) or glycoside ester(s) or a mixture comprising one or more kaempferol glycoside(s) and/or glycoside ester(s), wherein the, one, two, or three or all kaempferol glycosides or glycoside ester(s) is/are selected from the group consisting of Kaempferol 3-O-sophoroside, Kaempferol 3-sophoroside-7-0-glucoside, kaempferol 3-0-(2"'-0-sinapoyl-p- sophoroside), kaempferol 3-O-sinapoyl-sophoroside 7-O-glucoside (K3OSS), Kaempferol 3-0-(2"'-0-sinapoyl-p-sophoroside)-7-0-glucoside, Kaempferol 3-O-(2"'- O-galloyl-p-sophoroside), Kaempferol 3-0-(2"'-0-coumaroyl-p-sophoroside)), Kaempferol 3-0-(2"'-0-caffeoyl-p-sophoroside), Kaempferol 3-0-(2"'-0-feruloyl-p- sophoroside, preferably wherein the, one, two, or all kaempferol glycoside or glycoside ester is selected from the group consisting of kaempferol glucosides or glucoside esters, particularly preferably selected from the group consisting of kaempferol 3-O-sin- apoyl-sophoroside 7-O-glucoside (K3OSS), kaempferol 3-O-sophoroside, and kaempferol 3-0-(2"'-0-sinapoyl-p-sophoroside)-7-0-glucoside.

3. Use according to claim 1 or 2, wherein the compound or, respectively, one compound in the mixture is eriodictyol.

4. Use according to claim 1 or 2, wherein the compound or, respectively, one compound in the mixture is hesperetin.

5. Use according to claim 1 or 2, wherein the compound or, respectively, one compound in the mixture is naringenin.

6. Use according to claim 1 or 2, wherein the compound or, respectively, one compound in the mixture is phloretin.

7. Use according to any one of the preceding claims, wherein the mass ratio of i) the compound(s), selected from the group consisting of eriodictyol, hesperetin, naringenin, phloretin and mixtures thereof, and ii) kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof is in a range of from 1 :5 to 20:1 , preferably in the range of from 1 :2.5 to 15:1 , preferably in the range of from 1 :1 to 10:1 , preferably in the range of from 2.5:1 to 7.5 to 1 .

8. Method for reducing or masking the bitter taste impression of kaempferol or a glycoside thereof ora glycoside esterthereof ora mixture thereof, comprising the following steps i) providing kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, ii) providing eriodictyol, hesperetin, naringenin, phloretin, or a mixture thereof, iii) mixing the compound or mixture provided in step i) with the compound or mixture provided in step ii).

9. Method according to claim 8, for reducing or masking the bitter taste impression of one or more kaempferol glycoside(s) or glycoside ester(s) or a mixture comprising one or more kaempferol glycoside(s) and/or glycoside ester(s), wherein the, one, two, or three or all kaempferol glycosides or glycoside ester(s) is/are selected from the group consisting of Kaempferol 3-O-sophoroside, Kaempferol 3-sophoroside-7-0-glucoside, kaempferol 3-0-(2"'-0-sinapoyl-p- sophoroside), kaempferol 3-O-sinapoyl-sophoroside 7-O-glucoside (K3OSS), Kaempferol 3-0-(2"'-0-sinapoyl-p-sophoroside)-7-0-glucoside, Kaempferol 3-O-(2"'- O-galloyl-p-sophoroside), Kaempferol 3-0-(2"'-0-coumaroyl-p-sophoroside)), Kaempferol 3-0-(2"'-0-caffeoyl-p-sophoroside), Kaempferol 3-0-(2"'-0-feruloyl-p- sophoroside, preferably wherein the, one, two, or all kaempferol glycoside or glycoside ester is selected from the group consisting of kaempferol glucosides or glucoside esters, particularly preferably selected from the group consisting of kaempferol 3-O-sin- apoyl-sophoroside 7-O-glucoside (K3OSS), kaempferol 3-O-sophoroside, and kaempferol 3-0-(2"'-0-sinapoyl-p-sophoroside)-7-0-glucoside.

10. Method according to claim 8 or 9, wherein in step ii) eriodictyol or a mixture comprising eriodictyol is provided.

11 . Method according to claim 8 or 9, wherein in step ii) hesperetin or a mixture comprising hesperetin is provided.

12. Method according to claim 8 or 9, wherein in step ii) naringenin or a mixture comprising naringenin is provided.

13. Method according to claim 8 or 9, wherein in step ii) phloretin or a mixture comprising phloretin is provided.

14. Method according to any one of claims 8 to 13, wherein the mass ratio of eriodictyol, hesperetin, naringenin, phloretin or a mixture thereof, as provided in step ii), and kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, as provided in step i), is in a range of from 1 :5 to 20:1 , preferably in the range of from 1 :2.5 to 15:1 , preferably in the range of from 1 :1 to 10:1 , preferably in the range of from 2.5:1 to 7.5 to 1 .

15. Composition comprising i) eriodictyol, hesperetin, naringenin, phloretin, or a mixture thereof, and ii) kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, preferably a rapeseed protein isolate comprising kaempferol or a glycoside thereof or a glycoside ester thereof or mixtures thereof, wherein the amount of eriodictyol, hesperetin, naringenin, phloretin, or a mixture thereof is sufficient to reduce or mask the bitter taste impression of kaempferol or a glycoside thereof or a mixture thereof.

16. Composition according to claim 15, wherein the mass ratio of i) eriodictyol, hesperetin, naringenin, phloretin or a mixture thereof, and ii) kaempferol or a glycoside thereof or a glycoside ester thereof or a mixture thereof in the composition is in a range of from 1 :5 to 20:1 , preferably in the range of from 1 :2.5 to 15:1 , preferably in the range of from 1 :1 to 10:1 , preferably in the range of from 2.5:1 to 7.5 to 1 .

17. Nutritional preparation, semi-finished preparation or pharmacological preparation comprising a composition according to claim 15 or 16.