BRPI0811269B1 - ADDITIVE, AND, ADDITIVE AGENT - Google Patents

Description

Technical Field The present invention relates to a sweetener and an additive agent for enhancing the body taste of a sweet substance that is used as an ingredient of the sweetener, and for enhancing the bitter taste typical of the substance. sweetener.

Prior Art In recent years, a high intensity sweetener has been used as a sweetener in a wide variety of the food field such as dietary foods because of its high sweetness magnification. In the beverage and sweet fields each consuming a special amount. While large in sweeteners, various high intensity sweeteners have been used such as lower calorie sweeteners such as a reduced calorie sweetener and a calorie free sweetener, and sugar free sweeteners. Of those, typical examples of high intensity sweetener to be used include aspartame (APM), sucralose, acesulfame-K (Ace-K), neotame, N-N-β- (3-hydroxy-4) 1-methyl ester -methoxy-phenyl) -propyl-α-aspartyl-L-phenyl-alanine (hereinafter abbreviated as "ANS9801"), saccharin, stevia, glycyrhizine, thaumatin and monelin.

However, in general, consumers have become familiar over the years with the sweet taste of sugar, fruit glucose syrup or the like, which has been used for food in the past, and consequently do not in many cases prefer the quality of sweet taste. of such a high intensity sweetener as described above. For example, APM has a usability effect such as a flavor enhancing effect and an effect to reduce the bitter taste or astringency of coexisting substances, while having a problem with its remaining strong sweet taste (remaining strong sweet taste). and storage stability such as heat resistance. Sucralose has a high stability, while it has a problem in its taste suppressing effect and intense sweet taste in remaining flavor, for example. Ace-K has a high stability, while it has an intense intermediate-taste to remaining sweet taste (intense sweet over remaining taste) and particularly has a strong bitter taste and astringency, so Ace-K is often used in a combination with other high intensity sweeteners.

Hitherto, the use of those high intensity sweeteners in combination is known as a technology for modifying the sweet taste quality of the above mentioned high intensity sweetener (Patent Document 1). In addition, the use of those high-intensity sweeteners in combination with other sweeteners such as trehalose and erythritol (Patent Document 2) or an α-glycosylated stevia extract (Patent Document 3) and use in combination with a dietary fiber. (Patent Document 4) have been reported. However, at present, they have not yet sufficiently satisfied the preferences of consumers who have become familiar with the quality of sweet sugar taste or glucose fructose syrup.

Meanwhile, the calcium receptor which is also called Calcium Sensitive Receptor (CaSR) is a receptor consisting of 1,078 amino acids, which is classified in class C of seven transmembrane receptors (G protein coupled receptor; GPCR). Calcium receptor gene cloning was reported in 1993 (Non-Patent Document 1), and it is known that calcium receptor causes various cellular responses via elevation of intracellular calcium level, etc. when being activated with calcium etc. The human calcium receptor nucleotide sequence is recorded under GenBank Accession Number NM_000388 and is well conserved among animals. The above-mentioned calcium receptor may act to promote or suppress biological functions. Therefore, at the moment, a therapeutic agent utilizing an activating effect on the calcium receptor and a therapeutic agent utilizing an inhibiting effect on the calcium receptor are suitably used in the treatment of neurological diseases, liver diseases, cardiovascular diseases, digestive system, and other diseases, depending on pathological conditions. For example, the calcium receptor has functions to detect increased blood parathyroid calcium levels, and to suppress parathyroid hormone (PTH) secretion to correct blood calcium levels. Therefore, a blood calcium lowering effect is expected for a calcium receptor activator. It has really been clarified that when the calcium receptor activator is used to treat secondary hyperparathyroidism in a hemodialysis patient, it lowers the PTH level without raising the calcium and phosphorus levels.

Since functional analysis of calcium receptor has been conducted primarily for calcium homeostasis, applied research has thus far mainly focused on bone diseases in which calcium regulation is involved. However, it has become clear that the calcium receptor is widely distributed in living bodies other than parathyroid and kidney from the results of gene expression analysis etc. (Non-Patent Documents 2 and 3), and their possibilities for being involved in various biological functions and causes of disease have been proposed. For example, it is conjectured that the calcium receptor is involved in the functions of the liver, heart, lung, gastrointestinal tract, lymphocyte, and pancreas. The inventors of the present invention have also confirmed that the calcium receptor is expressed in a wide variety of tissues in living bodies by analyzing RNAs extracted from rat tissues using RT-PCR. From the above points of view, the values of calcium receptor activators and inhibitors are presently rapidly increasing for applications.

In addition, in addition to calcium, cations such as a gadolinium cation, basic peptides such as polyarginine, polyamine such as spermine, amino acids such as phenylalanine, and so on have been reported as calcium receptor activators. Non-Patent 4) · Non-Patent Document 5 reports that glutathione (γ-Glu-Cys-Gly), a low molecular weight peptide, is a CaSR activator, but in no way mentions a possibility of CaSR being involved. at the taste reception.

However, it is not known that an amino acid or peptide having a particular structure is useful as a calcium receptor activator. Furthermore, although a peptide derivative that exhibits a sweet taste such as aspartame is known, it is not known that an amino acid or peptide having a calcium receptor activating effect can enhance a sweet taste of a sweet substance. It should be noted that ANS9801 and neotame are described in Patent Documents 5 and 6, respectively.

[Patent Document 1] JP 2005-304440 A [Patent Document 2] JP 2002-51723 A [Patent Document 3] JP 2002-34501 A [Patent Document 4] JP 2004-41118 A [Patent Document 5] US 6,548,096 [Patent Document 6] International Publication NO. W039979 pamphlet [Non-Patent Document 1] Nature, 1993, Vol. 366 (6455), p. 575-580 [Non-Patent Document 2] J. Endocrinol., 2000, Vol. 165 (2), p. 173-177 [Non-Patent Document 3] Eur. J. Pharmacol., 2002, Vol. 447 (2-3), p. 271-278 [Non-Patent Document 4] Cell Calcium, 2004, Vol. 35 (3), ρρ. 209-216 [Non-Patent Document 5] J. Biol. Chem., 2006, Vol. 281 (13), p. Problems to be solved by the invention. Objectives of the present invention are: 1) to provide a sweetener in which sweet taste quality, in particular body taste and bitter taste have improved, especially a high sweetener. intensity at which sweet taste quality, in particular body taste and bitter taste have improved; and 2) providing additive agent to enhance the body taste of a sweet substance that is used as an ingredient of the sweetener, and to enhance the bitter taste typical of the sweet substance.

Means for Solving Problems The inventors of the present invention have found various amino acids and peptides as compounds each having a calcium receptor activating effect in the process of screening for a calcium receptor activator, and have also found that amino acids and peptides can improve quality. of sweet taste of a high sweetness sweetener, having thus accomplished the present invention.

That is, the present invention is as follows: (1) a sweetener comprising a sweet substance and a compound having calcium receptor activating effect; (2) the sweetener according to (1) wherein the compound comprises one or more amino acid or peptide types selected from the group consisting of γ-Glu-X-Gly where X represents an amino acid or amino acid derivative, γ-Glu -Val-Y where Y represents an amino acid or amino acid derivative, γ-Glu-Ala, γ-Glu-Gly, γ-Glu-Cys, γ-Glu-Met, y-Glu-Thr, γ-Glu-Val , γ-Glu-Om, Asp-Gly, Cys-Gly, Cys-Met, Glu-Cys, Gly-Cys, Leu-Asp, D-Cys, y-Glu-Met (O), y-Glu-y- Glu-Val, γ-Glu-Val-NH2, γ-Glu-Val-ol, γ-Glu-Ser, γ-Glu-Tau, γ-Glu-Cys (S-Me) (0), γ-Glu- Leu, γ-Glu-Ile, γ-Glu-t-Leu and γ-Glu-Cys (S-Me); (3) the sweetener according to (2) wherein X represents Cys (SNO), Cys (S-ally), Gly, Cys (S-Me), Abu, t-Leu, Cie, Alb, Pen or Ser, and Y represents Gly, Val, Glu, Lys, Phe, Ser, Pro, Arg, Asp, Met, Thr, His, Om, Asn, Cys, Gin, GlyA or LacA; (4) the sweetener according to (2) or (3) wherein the amino acid or peptide is selected from γ-Glu-Val-Gly and γ-Glu-Abu-Gly; (5) the sweetener according to any one of (1) to (4) wherein the sweet substance comprises one or more substance types selected from the group consisting of aspartame, sucralose, acesulfame K, neotame, ANS9801, saccharin, stevia, glycyrhizine, thaumatin, monatin and monelin; (6) an additive agent comprising a compound having calcium receptor activating effect, which is added in a sweet substance to enhance a body taste of the sweet substance; and (7) an additive agent comprising a compound having calcium receptor activating effect which is added in a sweet substance to enhance a bitter taste of the sweet substance.

Brief Description of the Drawings FIG. 1 is a graph showing a calcium effect on a calcium receptor. Human calcium receptor cRNA was injected into Xenopus laevis oocyte by microinjection. Values of intracellular response currents flowing at the addition of a calcium chloride solution at an arbitrary concentration were recorded. The maximum values of intracellular currents were considered response current values. It was confirmed that no response was observed in oocytes injected with microinjection distilled water as a control. FIG. 2 is a graph showing an effect of an L-amino acid on a calcium receptor. Human calcium receptor cRNA was injected into the Xenopus laevis oocyte by microinjection. Values of intracellular response currents flowing at the time of addition of a 10 mM L-amino acid solution were recorded. The maximum values of intracellular currents were considered response current values. It was confirmed that no response was observed in oocytes injected with microinjection distilled water as a control. FIG. 3 is a graph showing an effect of a D-amino acid on a calcium receptor. Human calcium receptor cRNA was injected into the Xenopus laevis oocyte by microinjection. Values of intracellular response currents that flowed upon the addition of a 10 mM D-amino acid solution were recorded. The maximum values of intracellular currents were considered response current values. It was confirmed that no response was observed in oocytes injected with microinjection distilled water as a control. FIG. 4 is a graph showing an effect of a peptide on a calcium receptor. Human calcium receptor cRNA was injected into the Xenopus laevis oocyte by microinjection. Intracellular response current values that flow at the time of addition of a peptide solution at an arbitrary concentration. The maximum values of intracellular currents were considered response current values. It was confirmed that no response was observed in oocytes injected with microinjection distilled water as a control. FIG. 5 is a conceptual diagram showing the intensity of a body taste of a sweet substance.

Best Mode for Carrying Out the Invention Hereinafter, the present invention is described in detail.

One embodiment of the present invention relates to a sweetener containing a sweet substance and a compound, preferably an amino acid or a peptide, having calcium receptor activating effect. Further, another embodiment of the present invention relates to an additive agent containing a compound having a calcium receptor activating effect, which is added in a sweet substance for the purpose of enhancing the body taste and bitter taste of the sweet substance. Furthermore, still another embodiment of the present invention relates to a sweetener which is obtained by the addition of the additive agent containing a compound having calcium receptor activating effect.

First, the amino acid or peptide having calcium receptor activating effect is described.

The inventors of the present invention have found that the amino acid or peptide having a calcium receptor activating effect can impart kokumi to a food. Furthermore, the inventors of the present invention have found that the amino acid or peptide can enhance the body taste and bitterness of the sweet substance. It should be noted that the term "kokumi" means a taste that cannot be expressed by the five basic flavors, which are sweet taste, salty taste, acid taste, bitter taste, and umami, and it means a taste in which both marginal flavors of the flavors. basic, such as thickness, growth (mouth filling), continuity, and harmony as basic flavors are intensified.

Meanwhile, the term "body taste" of the sweet substance as used herein means that the sweet substance has harmony and taste mainly in initial flavor and intermediate taste when eating the sweet substance. The phrase "improvement of body taste" refers to the intensification of body taste. The sense of taste varies with the course of time after the onset of eating, and is sequentially called immediately after the onset of eating, initial taste, intermediate taste and remaining taste. Those are relative notions, and typically, initial taste, intermediate taste, and remaining taste refer to flavors that are left 0 to 2 seconds, 3 to 4 seconds, and 5 seconds or later after the start of eating, respectively. . FIG. 5 shows a conceptual diagram of body taste, wherein the ordinate axis represents the intensity of the taste, and the abscissa axis represents the time after the onset of eating. However, FIG. 5 is a conceptual diagram and does not show absolute values. The term "calcium receptor" as used herein refers to a receptor which is called Calcium Sensitive Receptor (CaSR) and belongs to the class C of seven transmembrane receptors. The term "calcium receptor activator" refers to a substance that binds to the above-mentioned calcium receptor to activate the calcium receptor. In addition, the phrase "to activate a calcium receptor" as used herein means that a binder binds to a calcium receptor to activate a guanine nucleotide linker protein, thereby to transmit a signal. In addition, the term "calcium receptor activity" means that the calcium receptor transmits the signal.

Here, each amino acid and amino acids forming each peptide are L-amino acids unless otherwise indicated. <1> Compound having a calcium receptor activating effect The compound having a calcium receptor activating effect may be an amino acid, a peptide or derivatives thereof, or various low molecular weight compounds provided the compound has a sweetening quality enhancing effect of the sweet substance. The compound may also be a novel compound obtained by screening. For example, the compound may be obtained by reacting a calcium receptor with a test substance, and detecting calcium receptor activity. It is preferable to confirm that the obtained compound has a sweetening quality enhancing effect of the sweet substance, in particular of the high intensity sweetener.

Hereinafter, a compound screening method having a calcium receptor activating effect is specifically described, but is not limited to the following steps of: 1) measuring a calcium receptor activity by adding a test substance to a system measuring calcium receptor activity to measure calcium receptor activity; 2) comparing calcium receptor activity when the test substance is added with a calcium receptor activity when the test substance is not added; and 3) selecting the test substance exhibiting a calcium receptor activating effect at the time of addition of the test substance. Calcium receptor activity is measured by a measuring system using cells that express calcium receptors, for example.

The cells mentioned above may be cells endogenously expressing calcium receptors, or they may be recombinant cells into which an exogenous calcium receptor gene is introduced. The calcium receptor activity measuring system described above may be used without any particular limitation provided that when a calcium receptor-specific extracellular ligand (activator) is added to the above-mentioned cells expressing calcium receptors, the system The measuring device can detect the binding (reaction) between the activator and the calcium receptor, or it can respond to the binding (reaction) between the activator and the calcium receptor to thereby transmit a detectable signal into the cells. When calcium receptor activity is detected by reaction with the test substance, it is determined that the test substance has a calcium receptor stimulation activity, and is a substance that can enhance the sweet taste quality of the sweet substance. .

Meanwhile, a sweetening quality enhancing effect of the sweet substance can be confirmed by a method such as a human taste test. In addition, the amino acid and peptide to be used as a test substance are not particularly limited, the peptide is preferably a peptide consisting of 2 to 10 amino acid residues, or a derivative thereof, and more preferably a peptide consisting of 2 or 3. amino acid residues or a derivative thereof. In addition, the amino acid residue at the N-terminus of the peptide is preferably γ-glutamic acid.

Examples of the above mentioned calcium receptor include an animal calcium receptor derivative such as a mouse, rat and dog as well as human, the origin of the calcium receptor is not particularly limited.

As described above, calcium receptor activity can be confirmed by the use of living cells expressing a calcium receptor or fragment thereof, cell membranes expressing a calcium receptor or fragment thereof, an in vitro system containing a protein of a receptor of calcium. calcium or its fragment or the like.

An example using live cells is described below. However, confirmation of calcium receptor activity is not limited to this example.

A calcium receptor is expressed on cultured cells such as those from Xenopus laevis oocytes, hamster ovary cells, and human fetal kidney cells. The calcium receptor may be expressed by cloning a calcium receptor gene into a plasmid carrying a foreign gene and introducing the plasmid or cRNA obtained by using the plasmid as a template in cells. To detect the reaction, an electrophysiological technique and a fluorescent indicator indicating an increase in intracellular calcium level can be used.

Calcium receptor expression is first confirmed based on the response to calcium or a specific activator. Oocytes are used in which intracellular current is observed in response to 5 mM calcium, or cultured cells in which fluorescence of the fluorescent indicator reagent is observed in response to 5 mM calcium. Dependence on calcium concentration is determined by changing calcium concentration. Then, a test substance such as a peptide is prepared at a concentration of about 1 μΜ to 1 mM, and added to cultured oocytes or cells, and the calcium receptor activity of the above-mentioned peptide or the like is determined.

Examples of the compound to be used in the present invention (hereinafter referred to as the "compound of the present invention") include various amino acids, peptides, or derivatives thereof, or various low molecular weight compounds, each having a receptor activating effect. calcium (hereinafter, when "amino acid" or "peptide" is simply used, the amino acid and peptide sometimes mean both an amino acid and an amino acid derivative, and both a peptide and a peptide derivative, respectively). The amino acid or peptide having a calcium receptor activating effect to be used in the present invention may be an amino acid or peptide having an effect on enhancing the body taste and / or bitterness of the sweet substance when the amino acid or peptide is eaten. with the sweet substance. Examples of such an amino acid or such peptide include γ-Glu-X-Gly where X represents an amino acid or amino acid derivative, γ-Glu-Val-Y where Y represents an amino acid or amino acid derivative, γ-Glu- Wing, y-Glu-Gly, γ-Glu-Cys, γ-Glu-Met, γ-Glu-Thr, γ-Glu-Val, γ-Glu-Om, Asp-Gly, Cys-Gly, Cys-Met, Glu-Cys, Gly-Cys, Leu-Asp, D-Cys, y-Glu-Met (O), y-Glu-y-Glu-Val, γ-Glu-Val-NH2, γ-Glu-Val-ol , γ-Glu-Ser, γ-Glu-Tau, y-Glu-Cys (S-Me) (0), γ-Glu-Leu, γ-Glu-Ile, γ-Glu-t-Leu and y-Glu -Cys (S-Me) (hereinafter also referred to as the "peptide or the like of the present invention"). In the present invention, one type of such peptides or the like may be used, or two or more types thereof may be used in combination.

In addition, the peptide may be a peptide derivative having a structure of γ-G1u-X-OCH (Z) CO2 H (X represents an amino acid or amino acid derivative, and Z represents H (a hydrogen atom) or CH3 (a In addition, the peptide may be a compound in which Y represents GlyA or LacA in the formula γ-Glu-Val-Y. Its preferred specific examples include γ-Glu-Val-GlyA, γ-Glu-tLeu-GlyA, γ-Glu-Abu-GlyA, γ-Glu-Val-LacA, γ-Glu-tLeu-LacA, and γ-Glu-Abu-LacA It should be noted that GlyA represents glycolic acid and LacA represents lactic acid. Being one of S-lactic acid and R-lactic acid, S-lactic acid is preferred Structural formulas of those compounds are described below.

Here, examples of the amino acid include: a neutral amino acid such as Gly, Ala, Val, Leu, Ile, Ser, Thr, Cys, Met, Asn, Gin, Pro, Hyp, and t-Leu, an acidic amino acid such as Asp and Glu; a basic amino acid such as Lys, Arg, and His; an aromatic amino acid such as Phe, Tyr, and Trp; homoserine, citrulline, omithine, α-amino-butyric acid, norvaline, norleucine, and taurine. The amino acid may also be a non-naturally occurring amino acid (non-protein constituent) such as tert-leucine, cycloleucine, α-amino isobutyric acid, and L-penicylamine. It should be noted that X in the γ-Glu-X-Gly peptide may be any amino acid or derivative thereof described above, and preferably a non-Cys amino acid or derivative thereof.

Here, abbreviations for amino acid residues mean the following amino acids. (1) Gly: Glycine (31) Aib: α-Amino-isobutyric acid (2-methyl-alanine) (32) Pen: L-Penicyl-amine In addition, examples of amino acid derivative include various derivatives of the above mentioned amino acids such as a special amino acid, an unnatural amino acid, an amino alcohol, and a substituted amino acid whose side chain such as the terminal carbonyl group, the terminal amino group, and the cisterne thiol group are substituted with various substituents. Examples of the substituent include an alkyl group, an acyl group, a hydroxyl group, an amino group, an alkyl amino group, a nitro group, a sulfonyl group, and various protecting groups, especially include: Arg (NO2): Ν-γ nitro arginine; Cys (SNO): S-nitro-cysteine; Cys (S-Me): S-methyl cysteine; Cys (S-allyl): S-allyl cysteine; Val-NH 2: valinamide; and Val-ol: valinol (2-amino-3-methyl-1-butanol).

It should be noted that here, y-Glu-Cys (SNO) -Gly has the following structural formula, and the "(0)" in the above formulas γ-Glu-Met (O) and γ-Glu-Cys (S-Me) ) (O) indicates a sulfoxide structure. Ο "(γ)" in the formula γ-Glu indicates that glutamic acid binds to another amino acid via the carboxyl group at the γ-position in glutamic acid. S-Nitrous Glutathione (GNSO) γ-Glu-X-Gly where X represents an amino acid or amino acid derivative, γ-Glu-Val-Y where Y represents an amino acid or amino acid derivative, γ-Glu-Ala, γ-Glu-Gly, γ-Glu-Cys, γ-Glu-Met, γ-Glu-Thr, γ-Glu-Val, γ-Glu-Om, Asp-Gly, Cys-Gly, Cys-Met, Glu- Cys, Gly-Cys, Leu-Asp, D-Cys, Y "Glu-Met (0), γ-Glu-y-Glu-Val, y-Glu-Val-NIL, y-Glu-Val-ol, y -Glu-Ser, γ-Glu-Tau, y-Glu-Cys (S-Me) (0), γ-Glu-Leu, y-Glu-Ile, γ-Glu-t-Leu and y-Glu-Cys (S-Me) each perfect the body taste and bitterness of the sweet substance.

Therefore, γ-Glu-X-Gly where X represents an amino acid or amino acid derivative, γ-Glu-Val-Y where Y represents an amino acid or amino acid derivative, γ-Glu-Ala, γ-Glu-Gly, γ-Glu-Cys, γ-Glu-Met, γ-Glu-Thr, γ-Glu-Val, γ-Glu-Om, Asp-Gly, Cys-Gly, Cys-Met, Glu-Cys, Gly-Cys, Leu-Asp, D-Cys, y-Glu-Met (O), γ-Glu-y-Glu-Val, y-Glu-Val-NH2, y-Glu-Val-ol, y-Glu-Ser, y -Glu-Tau, y-Glu-Cys (S-Me) (0), γ-Glu-Leu, y-Glu-Ile, γ-Glu-t-Leu and γ-Glu-Cys (S-Me) ( hereinafter also called "peptide and amino acid to be used in the present invention") may be used as an additive agent for a sweet substance which is added to a sweet substance to enhance the body taste and bitterness of the sweet substance. The compound to be used in the present invention may be used alone or as a mixture of two or more arbitrary types. Of those, a compound having the following structural formula: γ-Glu-X-Gly where X represents Cys (SNO), Cys (S-ally), Gly, Cys (S-Me), Abu, t-Leu, Cie, Aib , Pen or Ser, or γ-Glu-Val-Y where Y represents Gly, Go, Glu, Lys, Phe, Ser, Pro, Arg, Asp, Met, Thr, His, Om, Asn, Cys, Gin, GlyA or LacA is preferably used.

Of those compounds, preferred compounds are y-Glu-Val-Gly and y-Glu-Abu-Gly.

As the above mentioned compound to be used in the present invention, a compound that is commercially available may be used. Further, when the compound is a peptide, the peptide may be obtained by the appropriate use of a known technique such as (1) a chemical peptide synthesis method, or (2) a peptide synthesis method by an enzymatic reaction. Since numerous amino acid residues contained in the peptide to be used in the present invention are comparatively small as 2 or 3 residues, a method of chemical synthesis of the peptide is convenient. When chemically synthesized, the oligopeptide may be synthesized or semi-synthesized by use of a peptide synthesizer. An example of the chemical peptide synthesis method includes a solid phase peptide synthesis method. The synthesized peptide as described above may be purified by standard means such as ion exchange chromatography, high performance reverse phase liquid chromatography or affinity chromatography. Such a method of peptide solid phase peptide synthesis and subsequent purification of the peptide are well known in the art.

In addition, the peptide to be used in the present invention may also be produced by an enzymatic reaction. For example, the method described in International Publication pamphlet WO 2004/011653 may be used. That is, the peptide may also be produced by: reacting an amino acid or dipeptide in which the carboxyl terminus is esterified or amidated with an amino acid having a free amino group (for example, an amino acid in which the carboxyl group is protected) in the presence of a peptide producing enzyme; and purifying the dipeptide or tripeptide produced. Examples of the peptide-producing enzyme include a microorganism culture having an ability to produce a peptide, separate microbial cells from the culture, or a processed microorganism cell product, or a microorganism-derived peptide-producing enzyme.

It should be noted that the peptide to be used in the present invention is not only produced by such an enzymatic method or a chemical synthesis method as mentioned above, but may also exist, for example, in a plant such as a vegetable or fruit, a microorganism such as a yeast, and a yeast extract. When the peptide exists in natural products, the peptide that is extracted from those natural products may be used.

In addition, the peptide need not be isolated prior to use, and may be used as a fraction containing the peptide of the present invention in a large amount.

Examples of the low molecular weight compound to be used in the present invention include cinacalcet ((R) -N- (3- (3- (trifluoromethyl) phenyl) propyl) -1- (1-naphthyl) ethyl amine) and an analogous compound thereof. Examples of the cinacalcet analog compound include the compound represented by the following chemical formula (1) ((R) -N - [(4-ethoxy-3-methylphenyl) methyl] -1- (1-naphthyl) ethyl amine)) or the compound represented by the following chemical formula (2) ((R) -N- (3-phenyl-prop-2-enyl) -1- (3-methoxy-phenyl) -ethylamine). Those compounds may be synthesized by such a known method as described in US Patent No. 6,211,244, for example. In addition, commercially available products may also be used.

(D (2) The compound to be used in the present invention also includes the form of a salt When the peptide and amino acid to be used in the present invention are in the form of a salt, the salt may be a pharmacologically acceptable salt. of a salt with an acid group such as caboxyl group in the formula include: an ammonium salt, an alkali metal salt such as sodium and potassium, an alkaline earth metal salt such as calcium and magnesium, an aluminum salt a zinc salt; a salt with an organic amine such as triethylamine, ethanolamine, morpholine, pyrrolidine, piperidine, piperazine, and dicyclohexylamine; and a salt with a basic amino acid such as arginine and lysine. Examples of a salt with a basic group in the case where the basic group exists in the formula include: a salt with an inorganic acid such as hydrochloric acid, sulfuric acid, phosphoric acid, nitric acid, and hydrobromic acid; a salt with an organic carboxylic acid as is acetic acid, citric acid, benzoic acid, maleic acid, fumaric acid, tartaric acid, succinic acid, tannic acid, butyric acid, benzoic acid, pamoic acid, enantic acid, decanoic acid, theoic acid, salicylic acid, lactic acid, oxalic acid mandelic acid and malic acid; and a salt with an organic sulfonic acid such as methanesulfonic acid, benzene sulfonic acid, and p-toluenesulfonic acid. <2> Additive and Sweetening Agent of the Present Invention The compound to be used in the present invention, preferably the peptide and amino acid, may be used as an additive agent which is added in a sweet substance for the purpose of enhancing the body taste of the substance. candy. The peptide and amino acid to be used in the present invention may also be used as an additive agent which is added to a sweet substance for the purpose of enhancing the sweetness of the sweet substance. The compound may be used alone or as a mixture of two or more types. The compound to be used in the present invention may also be mixed with a sweet substance to form a sweetener. The additive agent of the present invention may consist of only one or two or more types of compounds chosen from the above-mentioned compound to be used in the present invention. In addition, the additive agent may be formed by the optional addition of other compounds or various additives each having a sweetening quality enhancing effect of the sweetener. The amount of additive that is added to the sweet substance is not particularly limited as long as the amount improves the sweetness quality of the sweet substance, in particular the body taste. Specific amount of the present invention is, for example, 0.000001% to 99.9999 wt%, preferably 0.00001% to 99.999 wt%, and more preferably 0.0001% to 99.99 wt% with respect to the present invention. high intensity sweetener.

It should be noted that the above description is a description of the mixing ratio with respect to the high intensity sweetener, and the amount of use of the compound of the present invention in a food or beverage is 1 ppb to 99.9% by weight, preferably 10 ppb to 10 wt%, and more preferably 1 ppm to 1 wt%. The sweetener of the present invention contains the sweet substance and the compound to be used in the present invention. The compound may be used alone or as a mixture of two or more types.

As the sweet substance, a high intensity sweetener is preferably used, and examples thereof include aspartame, sucralose, acesulfame K, neotame, ANS9801, saccharin, stevia, glycyrhyrin, thaumatin, monatin and monelin. Of those, aspartame, sucralose and acesulfame K are preferably used. The sweet substance can be used alone or as a mixture of two or more types. It is well known that mixing multiple sweetening substances can enhance the body taste of each of the sweet substances. Even in such a case, the combined use of the peptide or amino acid to be used in the present invention with the sweet substance may further enhance body taste. Examples of the preferred combination of sweet substances include a combination of two optionally selected types of aspartame, sucralose and acesulfame-K, or a combination of those three types.

In addition, an example of the preferred combination includes a combination of aspartame and γ-Glu-Val-Gly which, as considered, has a high effect on the present invention. Specifically, this may be mixed so that the proportion of γ-Glu-Val-Gly by weight may be 0.1 to 500 parts by weight with respect to 100 parts by weight of aspartame. Of course, this is merely an example of mixing examples, and there is no need to be restricted to the aforementioned ratio. The sweetener of the present invention may naturally be mixed with common sweeteners such as sucrose, glucose, fructose and sugar alcohols (e.g. erythritol and maltitol). Examples of the sweetener form of the present invention include a powder, a granule and a liquid, and physical properties are not limited.

Examples Hereinafter, the present invention is more specifically described with reference to the examples. However, the scope of the present invention is not limited to these examples.

[Reference Example 1] Calcium Receptor Gene (cRNA) Preparation The calcium receptor gene gene was prepared as follows. Based on the NCBI-recorded DNA sequence (calcium receptor: NM_000388), synthetic oligo DNAs (forward primer (SEQ ID NO: 1) and reverse primer (SEQID NO: 2)) used for PCR were synthesized. PCR was performed by using the primers and Pfu ultra DMA Polymerase (manufactured by Stratagene) under the following conditions by using human kidney cDNA (manufactured by Clontech) as a material. After a reaction at 94 ° C for 3 minutes, a reaction cycle at 94 ° C for 30 seconds, 55 ° C for 30 seconds, and 72 ° C for 2 minutes was repeated 35 times, and then a reaction was performed at 72 ° C. ° C for 7 minutes. Whether amplification was achieved by PCR was confirmed by performing DNA agarose electrophoresis, staining the DNA with a DNA staining reagent, and subsequent ultraviolet irradiation. The chain lengths of the PCR products were conformed by comparison to DNA markers of known sizes simultaneously submitted to electrophoresis. Plasmid vector pBR322 was digested with restriction enzyme EcoRV (manufactured by Takara). The PCR-amplified gene fragment was ligated at the plasmid dividing site using the Binding Kit (manufactured by Promega). The Escherichia coli DH5a strain was transformed with each ligation reaction solution, and the transformant hosting the plasmid into which the PCR amplification product was cloned was selected. The PCR amplification product was confirmed by DNA base sequence analysis. By using the recombinant plasmid as a template together with a cDNA preparation kit (manufactured by Ambion), calcium receptor gene cRNA was prepared.

Reference Example 21 Preparation of Multiple Samples As a L-amino acid sample, 23 special grade amino acid types were used including alanine, arginine, asparagine, aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine, isoleucine, leucine, lysine , methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, valine, omithine, and taurine (all of these from Ajinomoto Co., Inc.), and hydroxy proline (Nacarai Tesque, Inc.). Like D-Cys and D-Trp (Nacarai Tesque, Inc.) and calcium chloride, those of special grade were used.

In addition, as a peptide sample, γ-Glu-Cys-Gly (Sigma Aldrich Japan KK), y-Glu-Cys (SNO) -Gly (Dojindo Laboratories), γ-Glu-Ala (Bachem Feinchemikalien AG), γ were used. -Glu-Gly (Bachem Feinchemikalien AG), γ-Glu-Cys (Sigma Aldrich Japan KK), γ-Glu-Met (Bachem Feinchemikalien AG), γ-Glu-Abu-Gly (Abu: a-amino-butyric acid, Bachem Feinchemikalien AG), γ-Glu-Thr (Kokusan Chemical Co., Ltd.), γ-Glu-Val (Kokusan Chemical Co., Ltd.), γ-Glu-Leu (ordered synthesis product), γ-Glu -Ile (order synthesis product), γ-Glu-Om (Kokusan Chemical Co., Ltd.), Asp-Gly (order synthesis product), Cys-Gly (order synthesis product), Cys-Met (order synthesis product) synthesis ordered), Glu-Cys (ordered synthesis product), Gly-Cys (ordered synthesis product), Leu-Asp (customs synthesis product), γ-Glu-Val-Val (ordered synthesis product), γ-Glu- Val-Glu (ordered synthesis product), γ-Glu-Val-Lys (ordered synthesis product), γ-Glu-y-Glu-Val (order synthesis product), γ-Glu-Gly-Gly (order synthesis product), γ-Glu-Val-Phe (order synthesis product), γ-Glu-Val-Ser (order synthesis product), γ -Glu-Val-Pro (order synthesis product), γ-Glu-Val-Arg (order synthesis product), γ-Glu-Val-Asp (order synthesis product), γ-Glu-Val-Met (product synthesis) Synthesis Ordered), γ-Glu-Val-Thr (Ordered Synthesis Product), γ-Glu-Val-His (Ordered Synthesis Product), γ-Glu-Val-Asn (Ordered Synthesis Product), γ-Glu Val-Gln (ordered synthesis product), γ-Glu-Val-Cys (ordered synthesis product), γ-Glu-Val-Om (ordered synthesis product) and γ-Glu-Ser-Gly (synthesis product) Glutamine and cysteine were prepared in use, and the other samples were stored at -20 ° C after preparation. Peptides having a purity of 90% or greater were used. For γ-Glu-Cys only, 80% or greater purity was used.

After dissolution of each sample, pH of the solution showing an acidic or alkaline pH was adjusted to approximately neutral pH by the use of NaOH or HCl. The composition of the solution used for dissolving amino acids and peptides, and the solution used for preparing Xenopus laevis oocytes, and for culturing the oocytes is as follows. 96 mM NaCl, 2 mM KCl, 1 mM MgCl 2, 1.8 mM CaCl 2, 5 mM Hepes, and pH 7.2. Reference Example 31 Synthesis of v-Glu-Val-Glv Boc-Val-OH (8.69 g, 40.0 mmol) and Gly-OBzl-HCl (8.07 g, 40.0 mmol) were dissolved in chloride methylene chloride (100 ml) and the solution was kept at 0 ° C. Triethylamine (6.13 ml, 44.0 mmol), HOBt (1-hydroxy-benzotriazole, 6.74 g, 44.0 mmol), and (1-ethyl-3- (3-hydroxy) WSC.HCl dimethylamino-propyl) carbodiimide, 8.44 g, 44.0 mmol) was added to the solution, and the mixture was stirred overnight at room temperature. The reaction solution was concentrated under reduced pressure, and the residue was dissolved in ethyl acetate (200 mL). The solution was washed with water (50 mL), 5% aqueous citric acid solution (50 mL x twice), saturated brine (50 mL), 5% aqueous sodium hydrogen carbonate solution (50 mL x twice), and saturated brine (50 ml). The organic layer was dried over anhydrous magnesium sulfate, magnesium sulfate was removed by filtration, and the filtrate was concentrated under reduced pressure. The residue was recrystallized from ethyl acetate / n-hexane to obtain Boc-Val-Gly-OBzl (13.2 g, 36.2 mmol) as a white crystal.

Boc-Val-Gly-OBzl (5.47 g, 15.0 mmol) was added in a 4 N HCl / dioxane solution (40 mL), and the mixture was stirred at room temperature for 50 minutes. Dioxane was removed by concentration under reduced pressure, n-hexane (30 mL) was added to the residue, and the mixture was concentrated under reduced pressure. The procedure was repeated 3 times to quantitatively obtain H-Val-Gly-OBzl.HCl. Δ-Val-Gly-OBzl-HCl and Z-Glu-OBzl (5.57 g, 15.0 mmol) described above were dissolved in methylene chloride (50 mL), and the solution was kept at 0 ° C. Triethylamine (2.30 ml, 16.5 mmol), HOBt (1-hydroxy-benzotriazole, 2.53 g, 16.5 mmol), and (1-ethyl-3- (3-hydroxy) WSC-HCl dimethylamino-propyl) carbodiimide, 3.16 g, 16.5 mmol) was added to the solution, and the mixture was stirred at room temperature overnight for 2 days. The reaction solution was concentrated under reduced pressure, and the residue was dissolved in heated ethyl acetate (1,500 ml). The solution was washed with water (200 mL), 5% aqueous citric acid solution (200 mL x twice), saturated brine (150 mL), 5% aqueous sodium hydrogen carbonate solution (200 mL x twice), and saturated brine (150 mL). The organic layer was dried over anhydrous magnesium sulfate, magnesium sulfate was removed by filtration, and the filtrate was concentrated under reduced pressure. The precipitated crystal was collected by filtration and dried under reduced pressure to obtain Z-Glu (Val-Gly-OBzl) -OBzl (6.51 g, 10.5 mmol) as a white crystal. Z-Glu (Val-Gly-OBzl) -OBzl (6.20 g, 10.03 mmol) described above was suspended in ethanol (200 mL), 10% palladium on carbon (1.50 g) was added to the suspension, and a reduction reaction was performed under a hydrogen atmosphere at 55 ° C for 5 hours. During the reaction, 100 ml in a total volume of water was gradually added. The catalyst was removed by filtration using a Kiriyama funnel, and the filtrate was concentrated under reduced pressure to half volume. The reaction solution was then filtered through a membrane filter, and the filtrate was concentrated under reduced pressure. The residue was dissolved in a small volume of water, and in the ethanol solution was added to precipitate a crystal, and the crystal was collected by filtration and dried under reduced pressure to obtain γ-Glu-Val-Gly as a white powder (2, 85 g, 9.40 mmol). ESI-MS: (M + H) + = 304.1. te-NMR (400 MHz, D 20) δ (ppm): 0.87 (3H, d, J = 6.8 Hz), 0.88 (3H, d, J = 6.8 Hz), 1.99- 2.0 9 (3H, m), 2.38-2.51 (2H, m) 3.72 (1H, t, δ, 35 Hz), 3.86 (1H, d, J-17.8 Hz) ), 3.80 (1H, d, J = 17.8 Hz), 4.07 (1H, d, J = 6.8 Hz).

[Reference Example 41 Synthesis of γ-Glu-Cvsi-MeVGlv rCvsi-Mei: Reduced S-methyl-cysteine Glutathione (15.0 g, 48.8 mmol) was added in water (45 ml) and sodium hydroxide (4 52 g, 2.2 equivalents, 107 mmol) was added portionwise while the mixture was bubbled with nitrogen. Methyl iodide (4.56 ml, 1.5 equivalents, 73 mmol) was added to the mixture, and the solution was sealed and stirred at room temperature for 2 hours. The reaction solution was adjusted to pH 2 to 3 with concentrated hydrochloric acid, and ethanol (150 mL) was added to the reaction solution, and stored overnight in a refrigerator. An oily product was separated from the supernatant was removed. When the remaining oily product was dissolved in water and ethanol was gradually added to the solution, a partially crystallized oily product was precipitated. The supernatant was again removed. The residue was dissolved in water (300 ml), adsorbed onto a column filled with a to ion exchange resin (Dowex 1-acetate, 400 ml), washed with water, and then eluted with 1 N aqueous acetic acid solution. The eluate was concentrated under reduced pressure, and re-precipitated in water / ethanol to obtain γ-Glu-Cys (S-Me) -Gly as a white powder (5.08 g, 15.8 mmol). FAB-MS: (M + H) + = 322. 1 H-NMR (400 MHz, D 20) δ (ppm): 2.14 (3H, s), 2.15-2.22 (2H, m), 2.50-2.58 (2H, m), 2.86 (1H, dd, J = 9.0 Hz, J = 14.0 Hz), 3.03 (1H, dd, J-5.0 Hz , J-14.0 Hz), 3.84 (1H, t, J = 6.5 Hz), 3.99 (2H, s), 4.59 (1H, dd, J = 5.0 Hz, J = 9.0 Hz).

[Reference Example 51 Synthesis of Other Peptides γ-Glu-Met (O), γ-Glu-Val-NH2, γ-Glu-Val-ol, γ-Glu-Ser, γ-Glu-Tau, y-Glu- Cys (S-Me) (0), γ-Glu-t-Leu, γ-Glu-Cys (S-allyl) -Gly, and γ-Glu-Cys (S-Me) were synthesized according to Examples 3 and 4.

[Reference Example 61 Evaluation of a Calcium Receptor Activating Effect For evaluation of the calcium receptor activating effect, a Ca concentration-dependent Cl ion ion current measurement method was used using a Xenopus laevis oocyte expression system. . If activator is added in Xenopus laevis oocytes in which the calcium receptor is expressed, intracellular Ca ions increase. Then the Ca ion concentration-dependent Cl channel opens, and the value of the intracellular current changes as an ionic current. By measuring the change in intracellular current value, it can be confirmed whether or not the calcium receptor activating effect is present.

Specifically, Xenopus laevis's abdomen was opened, and a portion of eggs were removed and then treated with a 1% collagenase solution at 20 ° C for 2 hours to obtain individual oocytes. Within each oocyte, 50 nl of 1 pg / μΐ receptor cRNA or 50 nl of sterilized oocyte water were injected using a glass microcapillary, and the oocytes were cultured at 18 ° C for 2 to 3 days. For the culture, a solution obtained by the addition of 2 mM pyruvic acid, 10 U / ml penicillin, and 10 µg / ml streptomycin in the solution mentioned in Reference Example 2 was used. After culture, a test solution was added to the oocytes. injected with cRNA or sterile water. Electrophysiological measurement was performed using a Geneclamp 500 amplifier (manufactured by Axon) and an AxoScope 9.0 recording computer program (manufactured by Axon). Oocytes were voltage-clamped at -70 mV by the double electrode voltage clamping method, and intracellular current via the Cl ion dependent on Ca ion concentration was measured. The maximum intracellular current value was considered as the value of the response current. Reference Example 71 Evaluation of calcium effect for calcium receptor activation The calcium receptor activating effect was evaluated using the method described in Reference Example 6. That is, oocytes injected with calcium receptor cRNA or sterile water were prepared, and voltage-clamped at -70 mV by the dual electrode voltage clamping method. For voltage-clamped oocytes, calcium was added (2 mM, 5 mM, 10 mM, and 20 mM), and then Ca ion concentration-dependent Cl response current was measured. FIG. 1 shows the results. From the results, it was confirmed that calcium receptor cRNA injected into the oocytes was functionally expressed. Furthermore, since oocytes injected with water did not respond even to a high calcium concentration, it was confirmed that the calcium receptor was not expressed in the oocytes themselves.

[Reference Example 81 Evaluation of the Effect of L-Amino Acids for Calcium Receptor Activation The Effect of L-Amino Acids for Calcium Receptor Activation was evaluated using the method described in Reference Example 6. That is, cRNA-injected oocytes Calcium receptor or sterile water were prepared, and voltage-clamped at -70 mV by the dual electrode voltage clamping method. In voltage-clamped oocytes, alanine (10 mM), arginine (10 mM), asparagine (10 mM), aspartic acid (10 mM), cysteine (10 mM), glutamine (10 mM), glycine (10 mM), histidine (10 mM), isoleucine (10 mM), leucine (10 mM), lysine (10 mM), methionine (10 mM), phenyl alanine (10 mM), proline (10 mM), serine (10 mM), threonine (10 mM), tryptophan (10 mM), tyrosine (10 mM), valine (10 mM), omithine (10 mM), taurine (10 mM), or hydroxy proline (10 mM) was added, and the current ion response concentration-dependent Cl response was measured. FIG. 2 shows the results. The results demonstrated that cysteine, histidine, phenyl alanine, tryptophan, and tyrosine each had a definite calcium receptor activating effect. The activating effect of the above mentioned amino acids has been reported in Proc. Natl. Acad. Know. USA, Apr. 25, 2000, 97 (9): 4814-9.

[Reference Example 91 Evaluation of the Effect of D-Cysteine for Calcium Receptor Activation The effect of D-cysteine for calcium receptor activation was evaluated using the method described in Reference Example 6. That is, cRNA-injected oocytes Calcium receptor or sterile water were prepared, and voltage-clamped at -70 mV by the double electrode voltage clamping method. In voltage-clamped oocytes, D-cysteine (10 mM), L-cysteine (10 mM), D-tryptophan (10 mM), or L-tryptophan (10 mM) were added, and current response was measured. Cl dependent on Ca ion concentration. FIG. 3 shows the results. From the results it is shown that D-cysteine had a definite calcium receptor activating effect.

["Reference Example 101 Evaluation of Peptide Effect for Calcium Receptor Activation The peptide effect for calcium receptor activation was evaluated using the method described in Reference Example 6. That is, cRNA-injected oocytes from Calcium or sterile water were prepared, and voltage-clamped at -70 mV by the double-electrode voltage clamping method.In voltage-clamped oocytes, γ-Glu-Cys-Gly (50 μΜ), y-Glu-Cys were added. (SNO) -Gly (50 μΜ), γ-Glu-Ala (50 μΜ), γ-Glu-Gly (500 μΜ), γ-Glu-Cys (50 μΜ), γ-Glu-Met (500 μΜ), γ-Glu-Thr (50 μΜ), γ-Glu-Val (50 μΜ), y-Glu-Om (500 μΜ), Asp-Gly (1 mM), Cys-Gly (1 mM), Cys-Met ( 1 mM), Glu-Cys (50 μΜ), Gly-Cys (500 μΜ), or Leu-Asp (1 mM), and current response of Cl ion dependent on Ca concentration was measured. From the results it is shown that the above-mentioned peptide had an effect on receptor activation. r of defined calcium.

[Reference Example 111 Evaluation of Calcium Receptor Activation Peptide Effect The peptide effect for calcium receptor activation was evaluated in the same manner as Reference Example 10. Each of the peptides shown in Table 1 was added in voltage-clamped oocytes at 1,000 μΜ, 300 μΜ, 100 μΜ, 30 μΜ, 10 μΜ, 3 μΜ, 1 μΜ, 0,3 μΜ, and 0,1 μΜ, and Ca ion concentration-dependent current response was measured. The lowest concentration at which current was detected was shown in Table 1 as the activity. From the results it is revealed that those 32 peptide types each had a calcium receptor activating effect.

Table 1 Reference Example 121 Peptide and amino acid kokumi activity to be used in the present invention Typical examples are selected from γ-Glu-X-Gly (where X represents Cys (SNO), Cys (S-allyl), Gly , Cys (S-Me), Abu, or Ser), γ-Glu -Val-Y (where Y represents Gly, Val, Glu, Lys, Phe, Ser, Pro, Arg, Asp, Met, Thr, His, Om , Asn, Cys, or Gin), γ-Glu-Ala, γ-Glu-Gly, γ-Glu-Cys, γ-Glu-Met, γ-Glu-Thr, γ-Glu-Val, γ-Glu -Om , Asp-Gly, Cys-Gly, Cys-Met, Glu-Cys, Gly-Cys, Leu-Asp, D-Cys, y-Glu-Met (O), and-Glu-y-Glu-Val, y- Glu-Val-NH2, γ-Glu-Val-ol, γ-Glu-Ser, γ-Glu-Tau, y-Glu-Cys (S-Me) (0), y-Glu-Leu, γ-Glu Ile, γ-Glu-t-Leu, and y-Glu-Cys (S-Me), each of which had a calcium receptor activating effect, and the presence or absence of kokumi-granting activity was examined by a sensory evaluation test. The sensory evaluation test was performed as follows. In distilled water containing sodium glutamate (0.05 g / dl), inosinic acid monophosphate (0.05 g / dl), and calcium chloride (1 mM) each of alliin (S-allyl cysteine sulfoxide: experiment (kokumi grant activity control), γ-Glu-Cys-Gly, γ-Glu-Cys, γ-Glu-Ala, and γ-Glu-Val were mixed as a sample in an amount of 0.2 g / dl , and the presence or absence of kokumi granting activity was determined. For a sample, which is acidic when the sample is dissolved, the sample pH was adjusted with NaOH to pH 6.8 to 7.2 before use. Table 2 shows the results.

Table 2 Calcium receptor promoter kokumi grant activity Reference Example 131 Peptide kokumi grant activity to be used in the present invention A peptide in which a calcium receptor activating effect was verified was examined for intensity of its grant activity. kokumi by a quantitative sensory assessment test. The quantitative sensory evaluation test was performed as follows. In distilled water containing sodium glutamate (0.05 g / dl), inosinic acid monophosphate (0.05 g / dl), and sodium chloride (0.5 g / dl) each of γ-Glu-Cys- Gly (glutathione), γ-Glu-Ala, γ-Glu-Met, and γ-Glu-Val were mixed as a sample in an amount of 0.1 g / dl, and the intensity of kokumi conceding activity was measured.

For a sample, which is acidic when dissolved, the pH of the sample was adjusted with NaOH to pH 6.8 to 7.2 before use. Due to the fact that it is known that glutathione can grant kokumi to a food, and thus, glutathione was used as a comparative control. The test was performed with n = 3 based on the following sensory evaluation scores: control (0 points) and glutathione addition (3 points). Table 3 shows the results. It should be noted that the term "initial and intermediate taste" is a collective term of an initial taste and an intermediate taste.

Table 3 Reference Example 141 Peptide kokumi-granting activity to be used in the present invention A peptide in which a calcium receptor activating effect was verified was examined for intensity of its kokumi-granting activity by a quantitative sensory evaluation test. The quantitative sensory evaluation test was performed as follows. In distilled water containing sodium glutamate (0.05 g / dl), inosinic acid monophosphate (0.05 g / dl), and sodium chloride (0.5 g / dl) each of γ-Glu-Cys- Gly (glutathione), γ-Glu-Cys, γ-Glu-Val, and γ-Glu-Val-Gly were mixed as a sample in an amount of 0.1 g / dl, or 0.01 g / dl as required. , and activity intensity of granting kokumi was measured. For a sample, which is acidic when dissolved, the pH of the sample was adjusted with NaOH to pH 6.8 to 7.2 before use. The test was performed with n = 5 based on the following sensory evaluation scores: control (0 points) and glutathione addition (3 points). Table 4 shows the results.

Table 4 * Unmeasurable: Kokumi granting activity is too strong to measure by sensory evaluation. Reference Example 151 Peptide kokumi-granting activity to be used in the present invention A peptide in which a calcium receptor activating effect was verified was examined for intensity of its kokumi-granting activity by a quantitative sensory evaluation test. The quantitative sensory evaluation test was performed as follows. In distilled water containing sodium glutamate (0.05 g / dl), inosinic acid monophosphate (0.05 g / dl), and sodium chloride (0.5g / dl) each of γ-Glu-Cys-Gly (glutathione), γ-Glu-Abu-Gly, and γ-Glu-Val-Gly were mixed as a sample in an amount of 0.1 g / dl or 0.01 g / dl, and the intensity of grant activity Kokumi was measured. For a sample, which is acidic when dissolved, the pH of the sample was adjusted with NaOH to pH 6.8 to 7.2 before use. The test was performed with n = 12 based on the following sensory evaluation scores: control (0 points) and glutathione addition (3 points). Table 5 shows the results.

Table 5 Example 11 Sweetness Activity of the Compound to be Used in the Present Invention A peptide in which a calcium receptor activating effect and a kokumi-granting activity were verified was examined for its activities (body taste enhancement and enhancement). bitterness) over typical high intensity sweeteners by a quantitative sensory evaluation test. The quantitative sensory evaluation test was performed as follows. As for the standard solutions of each of the sweeteners, the following compounds were used. The following standard solutions have been adjusted to have almost the same degree of sweetness between them.

It should be noted that an effect of peptide addition on each of the high intensity sweeteners was evaluated with reference to sucrose.

In distilled water containing each of the aforementioned sweeteners, γ-Glu-Cys-Gly (glutathione), γ-Glu-Val-Gly or γ-Glu-Abu-Gly was mixed as a sample at concentrations of 0.0001 to 1 g / dl, respectively, and then each degree of body flavor enhancement and bitterness enhancement over sweet taste was measured.

For a sample that has stronger acid properties than the standard solution not containing the sample as the sample is dissolved, the pH of the sample solution was adjusted with NaOH to reach a pH range ± 0.2 with respect to the pH of the solution. standard before use. Sensory evaluation for body taste enhancement was performed with n = 12 based on the following criteria: a control (obtained by adding distilled water only and without the addition of glutathione and / or the peptide of the present invention) was set to 0 point, and by using the control as a standard, weak: -2 points; slightly weak: -1 point; slightly strong: 1 point, and strong: 2 points were defined respectively. A score higher than 0 points means that the sample has a bitterness enhancing effect.

On the other hand, the sensory evaluation for the improvement of the bitter taste was performed with n = 12 based on the following criteria: a control (obtained by adding distilled water only and without adding glutathione and / or the peptide of the present invention) was set to 0 points, and by using the control as a default, weak: -2 points; slightly weak: -1 point; slightly strong: 1 point, and strong: 2 points were defined respectively. A score lower than 0 points means that the sample has a bitter taste enhancing effect.

As a result of the assessment, activities were widely exhibited at the addition concentrations mentioned above. Tables 6 to 8 each show the results at typical concentrations. As explained from the results in Tables 6, 7 and 8, it was confirmed that the peptides of the present invention each have a body flavor enhancing effect at a lower concentration than glutathione. Furthermore, it is also confirmed that the peptide of the present invention has a bitterness enhancing effect. In general, the addition of peptides provides improved body taste and a reduction in bitter taste, resulting in a sweet taste close to sucrose.

Table 6 Table 8 [Example 21 Activity of the compound to be used in the present invention on sweet taste (II) A peptide in which a calcium receptor activating effect and a kokumi granting activity were verified was examined for its effects (enhancement of body taste and bitterness enhancement) over products in which high intensity sweeteners are typically combined by a quantitative sensory evaluation test. The quantitative sensory evaluation test was performed as follows. As for the standard combination solution of each of the sweeteners, the following compounds were used. The following standard solutions have been adjusted to have almost the same degree of sweetness between them. i) Sucrose (5 g / dl) ii) APM (0.02 g / dl) + Ace-K (0.005 g / dl) [APM: Ace-K = 8: 2 as a sweetness ratio] iii) Sucralose (0.0058 g / dl) + Ace-K (0.0075 g / dl) [sucralose: Ace-K = 7: 3 as a sweetness ratio] iv) APM (0.0125 g / dl) + sucralose (0.0042 g / dl) [APM: sucralose = 1: 1 as a sweetness ratio] v) APM (0.0125 g / dl) + sucralose (0.0021 g / dl) + Ace- K (0.0063 g / dl) [APM: sucralose: Ace-K = 2: 1: 1 as a ratio of sweetness degrees] It should be noted that a peptide addition effect on each of the high intensity sweeteners was evaluated with reference to sucrose.

In distilled water containing each of the sweeteners, γ-Glu-Cys-Gly (glutathione), γ-Glu-Val-Gly or γ-Glu-Abu-Gly was mixed as a sample at concentrations of 0.0001 to 1 g / dl, respectively, and then each degree of perfection of body flavor and improvement of bitterness over sweet taste was measured. The effect of peptide addition on each of the high intensity sweeteners was evaluated with reference to sucrose.

For a sample that has stronger acidic properties than the standard solution not containing the sample as the sample is dissolved, the sample pH was adjusted with NaOH to reach a pH range ± 0.2 with respect to the pH of the standard solution before. of use. Sensory evaluation for body taste enhancement was performed at n = 12 based on the following criteria: a control (obtained by adding distilled water only and without the addition of glutathione and / or the peptide of the present invention) was set to 0 point, and by using the control as a standard, weak: -2 points; slightly weak: -1 point; slightly strong: 1 point, and strong: 2 points were defined respectively. A score higher than 0 points means that the sample has a body flavor enhancing effect.

On the other hand, the sensory evaluation for the improvement of the bitter taste was performed with n = 12 based on the following criteria: a control (obtained by adding distilled water only and without adding glutathione and / or the peptide of the present invention) was set to 0 points, and by using the control as a default, weak: -2 points; slightly weak: -1 point; slightly strong: 1 point, and strong: 2 points were defined respectively. A score lower than 0 points means that the sample has a bitterness enhancing effect.

As a result of the assessment, activities were widely exhibited at the addition concentrations mentioned above. Tables 9 to 12 each show the results at typical concentrations. As explained from the results in Tables 9 to 12, it was confirmed that the peptides of the present invention each have a body flavor enhancing effect at a lower concentration than glutathione. Furthermore, it is also confirmed that the peptide of the present invention has a bitterness enhancing effect. In general, the addition of peptides provides improved body taste and a reduction in bitter taste, resulting in a sweet taste close to sucrose.

Table 9 Table 10 Table 11 Tnhpla 1?

[Example 31 Sweet taste activity of the compound to be used in the present invention (ΊΙΡ A peptide in which a calcium receptor activating effect and a kokumi granting activity have been verified, and cinacalcet which is known to have a calcium receptor activating effect were examined for their activities (body taste enhancement and bitterness enhancement) on typical high-intensity sweeteners by a quantitative sensory evaluation test.The quantitative sensory evaluation test was performed as follows. the following compounds were used: The following standard solutions have been adjusted to have almost the same degree of sweetness between them.

It should be noted that a sample addition effect on each of the high intensity sweeteners was evaluated with reference to sucrose.

In distilled water containing each of the sweeteners, γ-Glu-Val-Gly or cinacalcet was mixed as a sample at concentrations of 0.0001 to 1 g / dl, respectively, and then each degree of body taste enhancement and perfection enhancement. bitterness over sweet taste was measured.

For a sample that has stronger acidic properties than the standard solution not containing the sample as the sample is dissolved, the sample pH was adjusted with NaOH to reach a pH range ± 0.2 with respect to the pH of the standard solution before. of use. Sensory evaluation for body taste improvement was performed with n = 12 based on the following criteria: a control (obtained by adding distilled water only and without adding a sample) was set to 0 points, and by using the control as a pattern, weak: -2 points; slightly weak: -1 point; slightly strong: 1 point, and strong: 2 points were defined respectively. A score higher than 0 points means that the sample has a body flavor enhancing effect.

On the other hand, the sensory evaluation for bitter taste improvement was performed with n = 12 based on the following criteria: a control (obtained by adding distilled water only and without adding a sample) was set to 0 points, and by using the control as a pattern, weak: -2 points; slightly weak: -1 point; slightly strong: 1 point, and strong: 2 points were defined respectively. A score lower than 0 points means that the sample has a bitterness enhancing effect.

As a result of the assessment, activities were widely exhibited at the addition concentrations mentioned above. Tables 13 to 15 each show the results at typical concentrations. As explained from the results in Tables 13 to 15, it is confirmed that γ-Glu-Val-Gly and cinacalcet each have a body flavor enhancing effect at almost the same concentration. Furthermore, it is also confirmed that the sample of the present invention has a bitterness enhancing effect. In general, addition of the sample provides improved body taste and a reduction in bitter taste, resulting in a sweet taste close to sucrose.

Table 13 Table 14 Table 15 Example 41 Sweet Taste (IV) activity of the compound to be used in the present invention A peptide in which a calcium receptor activating effect and a kokumi-granting activity have been verified, and cinacalcet which is known to have a Calcium receptor activating effects were examined for their effects (body taste enhancement and bitter taste enhancement) on products in which high intensity sweeteners are typically combined by a quantitative sensory evaluation test. The quantitative sensory evaluation test was performed as follows. As for the combination solution of each sweetener, the following compound was used. The following standard solutions have been adjusted to have almost the same degree of sweetness between them. i) Sucrose (5 g / dl) ii) APM (0.02 g / dl) + Ace-K (0.005 g / dl) [APM: Ace-K = 8: 2 as a sweetness ratio] iii) Sucralose (0.0058 g / dl) + Ace-K (0.0075 g / dl) [sucralose: Ace-K = 7: 3 as a sweetness ratio] iv) APM (0.0125 g / dl) + sucralose (0.0042 g / dl) [APM: sucralose = 1: 1 as a sweetness ratio] v) APM (0.0125 g / dl) + sucralose (0.0021 g / dl) + Ace- K (0.0063 g / dl) [APM: sucralose: Ace-K = 2: 1: 1 as a ratio of sweetness degrees] It should be noted that a peptide addition effect on each of the high intensity sweeteners was evaluated with reference to sucrose.

In distilled water containing each of the aforementioned sweeteners, γ-Glu-Val-Gly or cinacalcet was mixed as a sample at concentrations of 0.0001 to 1 g / dl, respectively, and then each degree of improvement in body taste and flavor. Improvement of bitter taste over sweet taste was measured. For one sample, the effect of sample addition on each of the high intensity sweeteners was evaluated with reference to sucrose.

For a sample that has stronger acidic properties than the standard solution not containing the sample as the sample is dissolved, the sample pH was adjusted with NaOH to reach a pH range ± 0.2 with respect to the pH of the standard solution before. of use. The sensory evaluation for body taste improvement was performed with n = 12 based on the following criteria: a control (obtained by adding distilled water only and without sample addition) was defined as 0 point, and by using the control as the one pattern, weak: -2 points; slightly weak: -1 point; slightly strong: 1 point, and strong: 2 points were defined respectively. A score higher than 0 points means that the sample has a body flavor enhancing effect.

On the other hand, the sensory evaluation for bitter taste improvement was performed with n = 12 based on the following criteria: a control (obtained by adding distilled water only and without adding the sample) was set to 0 points, and by use of control as a pattern, weak: -2 points; slightly weak: -1 point; slightly strong: 1 point, and strong: 2 points were defined respectively. A score lower than 0 points means that the sample has a bitterness enhancing effect.

As a result of the assessment, activities were widely exhibited at the addition concentrations mentioned above. Tables 16 to 19 each show the results at typical concentrations. As explained from the results in Tables 16 to 19, it is confirmed that γ-Glu-Val-Gly and cinacalcet each have a body flavor enhancing effect at almost the same concentration. Furthermore, it is also confirmed that the sample of the present invention has a bitterness enhancing effect. In general, addition of the sample provides improved body taste and a reduction in bitter taste, and results in a sweet taste close to sucrose.

Table 16 Table 18 Table 19 [Example 5] Activity of the compound to be used in the present invention in high intensity sweetener-containing beverage yogurt A peptide in which a calcium receptor activating effect and a kokumi-granting activity have been verified, and cinacalcet which is known to have a calcium receptor activating effect were examined for their activities (body taste enhancement and bitter taste enhancement) in a beverage yogurt containing a typical high intensity sweetener by a quantitative sensory evaluation test. The quantitative sensory evaluation test was performed as follows. As for a standard beverage yogurt solution, the following compounds were used. The following standard solutions have been adjusted to have almost the same degree of sweetness between them. i) Sucrose (5 g / dl) ii) APM (0.025 g / dl) It should be noted that a sample addition effect on each of the high intensity sweeteners was evaluated with reference to sucrose.

In a beverage yogurt containing each of the sweeteners, γ-Glu-Val-Gly or cinacalcet was mixed as a sample at concentrations of 0.0001 to 1 g / dl, respectively, and then each degree of improvement in body taste and Improvement of bitter taste over sweet taste was measured.

For a sample that has stronger acidic properties than the standard solution not containing the sample as the sample is dissolved, the sample pH was adjusted with NaOH to reach a pH range ± 0.2 with respect to the pH of the standard solution before. of use. The sensory evaluation for body taste improvement was performed with n = 12 based on the following criteria: a control (obtained by adding distilled water only and without sample addition) was defined as 0 point, and by using the control as the one pattern, weak: -2 points; slightly weak: -1 point; slightly strong: 1 point, and strong: 2 points were defined respectively. A score higher than 0 points means that the sample has a body flavor enhancing effect.

On the other hand, the sensory evaluation for bitter taste improvement was performed with n = 12 based on the following criteria: a control (obtained by adding distilled water only and without adding the sample) was set to 0 points, and by use of control as a pattern, weak: -2 points; slightly weak: -1 point; slightly strong: 1 point, and strong: 2 points were defined respectively. A score lower than 0 points means that the sample has a bitterness enhancing effect.

As a result of the assessment, activities were widely exhibited at the addition concentrations mentioned above. Table 20 shows the results at typical concentrations. As explained from the results in Table 20, it is confirmed that γ-Glu-Val-Gly and cinacalcet each have a body flavor enhancing effect at almost the same concentration. Furthermore, it is also confirmed that the sample of the present invention has a bitterness enhancing effect. In general, addition of the sample provides improved body taste and a reduction in bitter taste, and results in a sweet taste close to sucrose.

Table 20 Industrial Applicability The use of the sweetener, in particular, the high intensity sweetener such as aspartame in combination with the compound having a calcium receptor activating effect, preferably the amino acid or peptide, enhances the body taste or bitter taste of the sweetener. high intensity sweetener such as aspartame. The sweetener of the present invention having the improved body taste and the bitter taste of the sweet taste may be widely used for general foods such as beverages and fruit ice cream.

In addition, the compound having a calcium receptor activating effect may be widely used as an additive agent capable of enhancing the body taste and bitter taste of the sweetener, in particular of the high intensity sweetener.

Claims (9)

A sweetener comprising a sweet substance and a compound having a calcium receptor activating effect, wherein the compound comprises one or more amino acid or peptide types selected from the group consisting of γ-GIu-X-Gly where X represents an amino acid or amino acid derivative, γ-Glu-Val-Y where Y represents an amino acid or amino acid derivative, γ-Glu-Ala, y-Glu-Gly, y-Glu-Cys, γ-Glu-Met , γ-Glu-Thr, γ-Glu-Val, γ-GIu-Om, Asp-Gly, Cys-Met, GIu-Cys, Gly-Cys, Leu-Asp, D-Cys, γ-GIu-Met (O ), γ-Glu-y-Glu-Val, γ-Glu-Val-NH2, γ-Glu-Val-ol, y-Glu-Ser, γ-Glu-Tau, and-GIu-Cys (S-Me) (O), γ-Glu-Leu, γ-Glu-Ile, γ-Glu-t-Leu and γ-Glu-Cys (S-Me), where Orn is ornithine. Sweetener according to claim 1, characterized in that X represents Cys (SNO), Cys (S-altl), Gly, Cys (S-Me), Abu, t-Leu, Cie, Aib, Pen or Ser. And Y represents Gly, Val, Glu, Lys, Phe, Ser, Pro, Arg, Asp, Met, Thr, Hys, Om, Asn, Cys, Gin, Gly A or LacA, where Abu represents α-amino acid. -butyric, Cie represents cycloleucine, Aib represents Î ± -amino isobutyric acid, Pen represents L-penicylamine, Gly A represents glycolic acid, Om is ornithine and LacA represents larch acid. Sweetener according to claim 1, characterized in that the compound is selected from y-Glu-Val-Gly and y-GIu-Abu-Gly, wherein Abu represents α-amino-butyric acid. Sweetener according to any one of claims 1 to 3, characterized in that the sweet substance comprises one or more types of substance selected from the group consisting of aspartame, sucralose. acesulfame K, ncotame, N- [N- [3- (3-hydroxy-4-methoxy-phenyl) -propyl] -Î ± -aspartyl] -L-phenylalanine, saccharin, stevia, glycyrbizine 1-methyl ester , thaumatin, monatin and monelin. Additive agent to be added to a sweet substance to enhance a body taste of the sweet substance, comprising a compound having a calcium receptor activating effect, wherein the compound comprises one or more amino acid or peptide types selected from the sweetener. group consisting of γ-Glu-X-Gly where X represents an amino acid or amino acid derivative, γ-Glu-Val-Y where Y represents an amino acid or amino acid derivative, γ-Glu-Ala, γ-Glu-Gly , γ-Glu-Cys, γ-Glu-Met, γ-Glu-Thr, γ-Glu-Val, γ-Glu-Om, Asp-Gly, Cys-Met, Glu-Cys, Gly-Cys, Leu-Asp , D-Cys, γ-Glu-Met (O), γ-Glu-y-Glu-Val, γ-Glu-Val-NH2, γ-Glu-Val-ol, γ-Glu-Ser, γ-Glu- Tau, γ-Glu-Cys (S-Me) (O), γ-Glu-Leu, γ-Glu-Ile, γ-Glu-t-Leu and y-Glu-Cys (S-Me), where Om It is omithine, and wherein the body taste of the sweet substance means harmony and richness in the initial taste and intermediate taste when eating the sweet substance. Additive agent to be added in a sweet substance to enhance a bitter taste of the sweet substance, comprising a compound having a calcium receptor activating effect, wherein the compound comprises one or more amino acid or peptide types selected from the sweetener. group consisting of γ-Glu-X-Gly where X represents an amino acid or amino acid derivative, γ-Glu-Val-Y where Y represents an amino acid or amino acid derivative, γ-Glu-Ala, γ-Glu-Gly , γ-Glu-Cys, γ-Glu-Met, γ-Glu-Thr, γ-Glu-Val, γ-Glu-Om, Asp-Gly, Cys-Met, Glu-Cys, Gly-Cys, Leu-Asp , D-Cys, γ-Glu-Met (Ο), γ-Glu-y-Glu-Val, γ-Glu-Val-NH2, γ-Glu-Val-ol, γ-Glu-Ser, γ-Glu Tau, γ-Glu-Cys (S-Me) (O), γ-Glu-Leu, γ-Glu-Ile, γ-Glu-t-Leu and y-Glu-Cys (S-Me), where Om It is omitin. Additive according to Claim 5 or 6, characterized in that X represents Cys (SNO), Cys (S-ally), Gly, Cys (S-Me), Abu, t-Leu, Cie, Aib. , Pen or Ser, and Y represents Gly, Val, Glu, Lys, Phe, Ser, Pro, Arg, Asp, Met, Thr, His, Om, Asn, Cys, Gin, GlyA or LacA, where Abu represents acid a -amino-butyric, Cie represents cycloleucine, Aib represents α-amino isobutyric acid, Pen represents L-penicylamine, GlyA represents glycolic acid, Om is omithine and LacA represents lactic acid. Additive according to claim 5 or 6, characterized in that the compound is selected from γ-Glu-Val-Gly and γ-Glu-Abu-Gly, wherein Abu represents α-amino-butyric acid. Additive according to claim 5 or 6, characterized in that the sweet substance comprises one or more substance types selected from the group consisting of aspartame, sucralose, acesulfame K, neotame, N-methyl ester. [N- [3- (3-hydroxy-4-methoxy-phenyl) -propyl] -α-aspartyl] -L-phenylalanine, saccharin, stevia, glycyrhizine, thaumatin, monatin and monelin.