Novel isolated peptide, protein hydrolysate containing the isolated peptide, and uses thereof

Abstract

The present invention relates to an isolated peptide having an amino acid sequence selected from the group consisting of Pro-Val-Leu-Lys (SEQ ID NO: 1), Pro-Leu-Pro-Arg (SEQ ID NO: 2), Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Pro-Glu-Arg-Asn-Glu (SEQ ID NO: 4), Pro-Gln-Pro-Glu-Arg (SEQ ID NO: 5), Glu-Phe-Asp-Glu-Lys-Pro-Ala-Asp (SEQ ID NO: 6), Glu-Asp-Ala-Pro-Val-Leu-His (SEQ ID NO: 7), Glu-Phe-Asp-Ala-Arg-Pro-Thr (SEQ ID NO: 8), Glu-Glu-Thr-Pro-Ser-His-Asp (SEQ ID NO: 9), Glu-Glu-Val-Glu-Glu-Glu-Glu-Val-Glu (SEQ ID NO: 10), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Glu-Asn-Ala-Pro-Asp-Gln-Lys (SEQ ID NO: 12), Glu-Asp-Met-Ala-Pro (SEQ ID NO: 13), Pro-Ile-Asn-Asp-Asn (SEQ ID NO: 14), Pro-Thr-Asp-Leu (SEQ ID NO: 15), Pro-Val-Ala-Glu (SEQ ID NO: 16), Arg-Asn-Gly-Pro-Arg (SEQ ID NO: 17), Arg-Ala-Glu-Asp-Thr-Ala-Thr-Tyr-Tyr (SEQ ID NO: 18), Glu-Gly-Asp-Ser-Ala-Ala-Ile (SEQ ID NO: 19), Glu-Pro-Glu-Gly-Asp-Val-His-Gln (SEQ ID NO: 20), and Glu-Asp-Glu-Val-Leu-Ala-Thr-Pro (SEQ ID NO: 21), a protein hydrolysate containing the isolated peptide, and its use as a taste modifier in food preparations and pharmaceutical preparations.

Description

Technical Field

[0001] The present invention relates to a novel isolated peptide for use as a taste regulator in food preparations and pharmaceutical preparations, and a protein hydrolyzate containing the isolated peptide.

Background Art

[0002] The blood of livestock and poultry is a by-product obtained during the process of industrial slaughter. The blood of livestock and poultry, such as chicken blood, is produced in large quantities every year and wasted in many countries around the world despite its valuable proteins and nutrients' bioavailability. Therefore, the utilization of such blood is economically and environmentally advantageous. So far, in the food industry, the use of whole blood or proteins has been restricted due to unpleasant flavors and colors, so the utilization of livestock and poultry blood has been restricted regardless of cost efficiency and feeding purposes. To address these problems, several studies have been conducted to recycle by-products and waste obtained from poultry processing and the poultry industry. For example, chicken blood is used to make blood sausage, blood tofu, and blood pudding. Plasma can be used as a protein supplement that is an excellent source of trace minerals, or as a substitute for egg white in the bakery industry due to its excellent foaming ability (Jayathilakan et al., J Food Sci Technol, 2012, 49(3), 378 - 293).

[0003] To increase the use of chicken blood, the generation of taste-active peptides and taste-regulating peptides has been proposed. However, existing knowledge about the composition of chicken blood, such as which proteins are contained or the exact concentrations of some components, is limited. Furthermore, although many studies have been conducted to identify taste-active peptides and taste-regulating peptides in all foods, there is no research on the taste of chicken blood. Due to such limited knowledge and the complex matrix of chicken blood, it is difficult to identify taste-active peptides and taste-regulating peptides from the blood of livestock and poultry, such as chicken blood.

[0004] CN 108047313 A discloses a chicken blood antioxidant peptide and a fermentation preparation method. The antioxidant peptide has the amino acid sequence Thr-Ser-Phe-Gly-Asp-Ala-Val-Lys-Asn-Leu-Asp-Asn-Ile-Lys (SEQ ID NO: 1). This antioxidant peptide has high free radical scavenging activity, a high scavenging effect on DPPH free radicals, and high reducing ability. This method includes the steps of activation of culture, preparation of blood cell medium, inoculation fermentation, separation, and purification.

[0005] CN 108048518 A discloses a chicken blood antioxidant peptide and an enzymatic hydrolysis preparation method. The antioxidant peptide is the amino acid sequence Met-Gly-Gln-Lys-Asp-Ser-Tyr-Val-Gly-Asp-Glu-Ala-Gln-Ser-Lys-Arg-Gly-Ile-Leu-Thr (SEQ ID NO: 1), or Ala-Glu-Asp-Lys-Lys-Leu-Ile-Gln (SEQ ID NO: 2). These antioxidant peptides have high free radical scavenging activity, a high scavenging effect on DPPH free radicals and superoxide anion free radicals, and high reducing ability. This method includes the steps of preparation of blood cells, hemolysis, protein degradation, and ultrafiltration.

[0006] CN 101843289 A discloses a method for producing blood polypeptide protein powder from livestock and poultry blood. According to this method, the poorly absorbable and utilizable high molecular albumin in livestock and poultry blood is decomposed into low molecular albumin, polypeptides, and free amino acids, improving the taste and nutritional value of dried blood for feed.

[0007] Despite the development of products and methods for using the above-mentioned livestock and poultry blood, the use of livestock and poultry blood, especially peptides derived from livestock and poultry blood having specific amino acid sequences, has not been disclosed as taste activators and taste regulators. Summary of the Invention

[0008] The present invention has been invented in consideration of the above problems.

[0009] Accordingly, one object of the present invention is to provide a novel taste-active peptide and a mixture thereof for a taste regulator of food preparations and pharmaceutical preparations by using animal blood, particularly livestock and poultry blood, which is industrial waste discharged from a meat processing step.

[0010] Another object of the present invention is to provide a peptide derived from animal blood having a specific amino acid sequence, particularly a peptide derived from livestock and poultry blood. The peptide derived from animal blood of the present invention can be used as a taste-active agent that imparts taste-regulating effects such as a saltiness-reducing effect, a saltiness-enhancing effect, a umami effect, and a kokumi effect.

[0011] Still another object of the present invention is to provide a protein hydrolyzate containing a peptide derived from animal blood and a taste-regulating composition.

[0012] A further object of the present invention is to provide a method for regulating the taste of food preparations and pharmaceutical preparations by using a peptide derived from animal blood, a protein hydrolyzate, and a taste-regulating composition.

[0013] In one aspect, the present invention relates to an isolated peptide having an amino acid sequence selected from the group consisting of Pro-Val-Leu-Lys (SEQ ID NO: 1), Pro-Leu-Pro-Arg (SEQ ID NO: 2), Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Pro-Glu-Arg-Asn-Glu (SEQ ID NO: 4), Pro-Gln-Pro-Glu-Arg (SEQ ID NO: 5), Glu-Phe-Asp-Glu-Lys-Pro-Ala-Asp (SEQ ID NO: 6), Glu-Asp-Ala-Pro-Val-Leu-His (SEQ ID NO: 7), Glu-Phe-Asp-Ala-Arg-Pro-Thr (SEQ ID NO: 8), Glu-Glu-Thr-Pro-Ser-His-Asp (SEQ ID NO: 9), Glu-Glu-Val-Glu-Glu-Glu-Glu-Val-Glu (SEQ ID NO: 10), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Glu-Asn-Ala-Pro-Asp-Gln-Lys (SEQ ID NO: 12), Glu-Asp-Met-Ala-Pro (SEQ ID NO: 13), Pro-Ile-Asn-Asp-Asn (SEQ ID NO: 14), Pro-Thr-Asp-Leu (SEQ ID NO: 15), Pro-Val-Ala-Glu (SEQ ID NO: 16), Arg-Asn-Gly-Pro-Arg (SEQ ID NO: 17), Arg-Ala-Glu-Asp-Thr-Ala-Thr-Tyr-Tyr (SEQ ID NO: 18), Glu-Gly-Asp-Ser-Ala-Ala-Ile (SEQ ID NO: 19), Glu-Pro-Glu-Gly-Asp-Val-His-Gln (SEQ ID NO: 20), and Glu-Asp-Glu-Val-Leu-Ala-Thr-Pro (SEQ ID NO: 21).

[0014] In another aspect, the present invention relates to a protein hydrolysate comprising at least one peptide having an amino acid sequence selected from the group consisting of Pro-Val-Leu-Lys (SEQ ID NO: 1), Pro-Leu-Pro-Arg (SEQ ID NO: 2), Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Pro-Glu-Arg-Asn-Glu (SEQ ID NO: 4), Pro-Gln-Pro-Glu-Arg (SEQ ID NO: 5), Glu-Phe-Asp-Glu-Lys-Pro-Ala-Asp (SEQ ID NO: 6), Glu-Asp-Ala-Pro-Val-Leu-His (SEQ ID NO: 7), Glu-Phe-Asp-Ala-Arg-Pro-Thr (SEQ ID NO: 8), Glu-Glu-Thr-Pro-Ser-His-Asp (SEQ ID NO: 9), Glu-Glu-Val-Glu-Glu-Glu-Glu-Val-Glu (SEQ ID NO: 10), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Glu-Asn-Ala-Pro-Asp-Gln-Lys (SEQ ID NO: 12), Glu-Asp-Met-Ala-Pro (SEQ ID NO: 13), Pro-Ile-Asn-Asp-Asn (SEQ ID NO: 14), Pro-Thr-Asp-Leu (SEQ ID NO: 15), Pro-Val-Ala-Glu (SEQ ID NO: 16), Arg-Asn-Gly-Pro-Arg (SEQ ID NO: 17), Arg-Ala-Glu-Asp-Thr-Ala-Thr-Tyr-Tyr (SEQ ID NO: 18), Glu-Gly-Asp-Ser-Ala-Ala-Ile (SEQ ID NO: 19), Glu-Pro-Glu-Gly-Asp-Val-His-Gln (SEQ ID NO: 20), Glu-Asp-Glu-Val-Leu-Ala-Thr-Pro (SEQ ID NO: 21), and combinations thereof.

[0015] In another aspect, the present invention relates to a taste-modulating composition comprising the isolated peptide or protein hydrolysate as defined above.

[0016] In yet another aspect, the present invention relates to a food preparation or a pharmaceutical preparation comprising the isolated peptide, protein hydrolysate, or taste-modulating composition as defined above.

[0017] Furthermore, the present invention relates to a method for modulating the taste of a food preparation or a pharmaceutical preparation, comprising applying an effective amount of the isolated peptide, protein hydrolysate, or taste modulating composition as defined above to the food preparation or the pharmaceutical preparation.

Brief Description of the Drawings

[0018] The features of the present invention will be more clearly described by reading the following description of specific embodiments. The above examples are for the purpose of assisting understanding and are not for the purpose of limitation.

[0019]

Figure 1

Figure 2

Modes for Carrying Out the Invention

[0020] Unless otherwise specified, any aspect shown in this specification is also intended to include its application to other aspects of the present invention.

[0021] Unless otherwise specified, the technical terms and scientific terms used in this specification have definitions understood by those skilled in the art.

[0022] Throughout the present invention, the term "about" is used to indicate that any value shown or presented in this specification may vary or deviate. Such variation or deviation may be the result of errors in the equipment or method used to determine the value.

[0023] The terms "consist(s) of" and its variations, such as "consisting of" and "consisted of", "comprise(s)" and its variations, such as "comprising" and "comprised", "has / have / having", and "include(s)" and its variations, such as "including" and "included", are free-form verbs. For example, any method of "consisting of", "comprising", "having", or "including" one or more components or steps is not limited to only one or more components or steps, but also includes components or steps not mentioned.

[0024] When the terms "a", "an", and "the" are used in the singular form, they are intended to include the plural form of the noun as well, unless otherwise specified.

[0025] As used herein, the term "modulating taste" is synonymous with the terms "improving taste", "adjusting taste", "reducing taste", "increasing taste", and "changing taste", which can be used interchangeably. In the present invention, the terms "taste modulator" and "taste modulating composition" refer to agents and compositions that can impart one or more tastes, such as saltiness, sweetness, umami, savoriness, astringency, bitterness, and sourness, to a food preparation or a pharmaceutical preparation and can change the taste of the food preparation or the pharmaceutical preparation as desired. Further, modulation of taste can be obtained by reducing or enhancing one or more of these tastes.

[0026] As used herein, the term "food preparation" refers to all food and beverage products, compositions, and formulations suitable for human and animal consumption, including, for example, foods, beverages, nutritional supplements, snacks, desserts, and candies. Similarly, the term "pharmaceutical preparation" as used herein refers to all pharmaceutical products, compositions, and formulations suitable for consumption. Food preparations and pharmaceutical preparations can be in solid (such as powders, tablets, granules, beads, and capsules), semi-solid, gel, and liquid forms.

[0027] Any means, apparatus, method, material, or chemical substance referred to herein means a means, apparatus, method, material, or chemical substance generally used or practiced by those skilled in the art, unless otherwise specified.

[0028] All chemical substances, compounds, materials, components, and / or methods disclosed in the present invention, as well as the claims, are intended to cover aspects of the present invention obtained from any action, practice, modification, or change added to the factors without performing experiments significantly different from the present invention, and although not particularly indicated in the claims, according to the opinion of those skilled in the art, it is intended to obtain objects having characteristics, usefulness, and effects similar to those of the present invention. Therefore, objects equivalent or similar to the aspects of the present invention, including minor modifications or changes obvious to those skilled in the art, should be considered to be within the spirit, scope, and concept of the present invention.

[0029] The present invention will be described in more detail below.

[0030] The first aspect of the present invention is an isolated peptide having an amino acid sequence selected from the group consisting of Pro-Val-Leu-Lys (SEQ ID NO: 1), Pro-Leu-Pro-Arg (SEQ ID NO: 2), Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Pro-Glu-Arg-Asn-Glu (SEQ ID NO: 4), Pro-Gln-Pro-Glu-Arg (SEQ ID NO: 5), Glu-Phe-Asp-Glu-Lys-Pro-Ala-Asp (SEQ ID NO: 6), Glu-Asp-Ala-Pro-Val-Leu-His (SEQ ID NO: 7), Glu-Phe-Asp-Ala-Arg-Pro-Thr (SEQ ID NO: 8), Glu-Glu-Thr-Pro-Ser-His-Asp (SEQ ID NO: 9), Glu-Glu-Val-Glu-Glu-Glu-Glu-Val-Glu (SEQ ID NO: 10), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Glu-Asn-Ala-Pro-Asp-Gln-Lys (SEQ ID NO: 12), Glu-Asp-Met-Ala-Pro (SEQ ID NO: 13), Pro-Ile-Asn-Asp-Asn (SEQ ID NO: 14), Pro-Thr-Asp-Leu (SEQ ID NO: 15), Pro-Val-Ala-Glu (SEQ ID NO: 16), Arg-Asn-Gly-Pro-Arg (SEQ ID NO: 17), Arg-Ala-Glu-Asp-Thr-Ala-Thr-Tyr-Tyr (SEQ ID NO: 18), Glu-Gly-Asp-Ser-Ala-Ala-Ile (SEQ ID NO: 19), Glu-Pro-Glu-Gly-Asp-Val-His-Gln (SEQ ID NO: 20), and Glu-Asp-Glu-Val-Leu-Ala-Thr-Pro (SEQ ID NO: 21).

[0031] The isolated peptide of the present invention can impart at least one of a saltiness-reducing effect, a saltiness-enhancing effect, an umami effect, and a kokumi effect.

[0032] In a preferred embodiment, the isolated peptide conferring a saltiness-reducing effect has an amino acid sequence selected from the group consisting of Pro-Val-Leu-Lys (SEQ ID NO: 1), Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), and Glu-Asp-Met-Ala-Pro (SEQ ID NO: 13).

[0033] In another preferred embodiment, the isolated peptide conferring a saltiness-enhancing effect has an amino acid sequence selected from the group consisting of Glu-Phe-Asp-Glu-Lys-Pro-Ala-Asp (SEQ ID NO: 6), Glu-Phe-Asp-Ala-Arg-Pro-Thr (SEQ ID NO: 8), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Pro-Ile-Asn-Asp-Asn (SEQ ID NO: 14), Pro-Val-Ala-Glu (SEQ ID NO: 16), and Glu-Asp-Glu-Val-Leu-Ala-Thr-Pro (SEQ ID NO: 21).

[0034] In a more preferred embodiment, the isolated peptide imparting an umami effect has an amino acid sequence selected from the group consisting of Pro-Val-Leu-Lys (SEQ ID NO: 1), Pro-Leu-Pro-Arg (SEQ ID NO: 2), Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Pro-Glu-Arg-Asn-Glu (SEQ ID NO: 4), Pro-Gln-Pro-Glu-Arg (SEQ ID NO: 5), Glu-Phe-Asp-Glu-Lys-Pro-Ala-Asp (SEQ ID NO: 6), Glu-Asp-Ala-Pro-Val-Leu-His (SEQ ID NO: 7), Glu-Glu-Thr-Pro-Ser-His-Asp (SEQ ID NO: 9), Glu-Glu-Val-Glu-Glu-Glu-Glu-Val-Glu (SEQ ID NO: 10), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Glu-Asn-Ala-Pro-Asp-Gln-Lys (SEQ ID NO: 12), Pro-Ile-Asn-Asp-Asn (SEQ ID NO: 14), Arg-Asn-Gly-Pro-Arg (SEQ ID NO: 17), Arg-Ala-Glu-Asp-Thr-Ala-Thr-Tyr-Tyr (SEQ ID NO: 18), Glu-Gly-Asp-Ser-Ala-Ala-Ile (SEQ ID NO: 19), Glu-Pro-Glu-Gly-Asp-Val-His-Gln (SEQ ID NO: 20), and Glu-Asp-Glu-Val-Leu-Ala-Thr-Pro (SEQ ID NO: 21).

[0035] In a more preferred embodiment, the isolated peptide imparting umami taste has an amino acid sequence selected from the group consisting of Pro-Val-Leu-Lys (SEQ ID NO: 1), Pro-Leu-Pro-Arg (SEQ ID NO: 2), Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Pro-Glu-Arg-Asn-Glu (SEQ ID NO: 4), Pro-Gln-Pro-Glu-Arg (SEQ ID NO: 5), Glu-Asp-Ala-Pro-Val-Leu-His (SEQ ID NO: 7), Glu-Phe-Asp-Ala-Arg-Pro-Thr (SEQ ID NO: 8), Glu-Glu-Thr-Pro-Ser-His-Asp (SEQ ID NO: 9), Glu-Glu-Val-Glu-Glu-Glu-Glu-Val-Glu (SEQ ID NO: 10), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Glu-Asn-Ala-Pro-Asp-Gln-Lys (SEQ ID NO: 12), Glu-Asp-Met-Ala-Pro (SEQ ID NO: 13), Pro-Thr-Asp-Leu (SEQ ID NO: 15), Pro-Val-Ala-Glu (SEQ ID NO: 16), Arg-Asn-Gly-Pro-Arg (SEQ ID NO: 17), Arg-Ala-Glu-Asp-Thr-Ala-Thr-Tyr-Tyr (SEQ ID NO: 18), Glu-Gly-Asp-Ser-Ala-Ala-Ile (SEQ ID NO: 19), Glu-Pro-Glu-Gly-Asp-Val-His-Gln (SEQ ID NO: 20), and Glu-Asp-Glu-Val-Leu-Ala-Thr-Pro (SEQ ID NO: 21).

[0036] The isolated peptide of the present invention is obtained from or derived from enzymatic hydrolysis or fermentation. Specifically, the isolated peptide is obtained from or derived from enzymatic hydrolysis of blood of animals including, but not limited to, pigs, cows, and poultry (such as chickens, birds, turkeys, etc.). In a preferred embodiment, the blood of the animal useful in the present invention is poultry blood, more preferably chicken blood.

[0037] In an exemplary embodiment, the enzymatic hydrolysis for obtaining the isolated peptide of the present invention is carried out using a serine protease. Preferably, the serine protease is subtilisin.

[0038] The present invention also provides the use of the isolated peptide defined above as a taste modifier for food preparations or pharmaceutical preparations.

[0039] A second aspect of the present invention is a protein hydrolysate comprising at least one peptide having an amino acid sequence selected from the group consisting of Pro-Val-Leu-Lys (SEQ ID NO: 1), Pro-Leu-Pro-Arg (SEQ ID NO: 2), Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Pro-Glu-Arg-Asn-Glu (SEQ ID NO: 4), Pro-Gln-Pro-Glu-Arg (SEQ ID NO: 5), Glu-Phe-Asp-Glu-Lys-Pro-Ala-Asp (SEQ ID NO: 6), Glu-Asp-Ala-Pro-Val-Leu-His (SEQ ID NO: 7), Glu-Phe-Asp-Ala-Arg-Pro-Thr (SEQ ID NO: 8), Glu-Glu-Thr-Pro-Ser-His-Asp (SEQ ID NO: 9), Glu-Glu-Val-Glu-Glu-Glu-Glu-Val-Glu (SEQ ID NO: 10), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Glu-Asn-Ala-Pro-Asp-Gln-Lys (SEQ ID NO: 12), Glu-Asp-Met-Ala-Pro (SEQ ID NO: 13), Pro-Ile-Asn-Asp-Asn (SEQ ID NO: 14), Pro-Thr-Asp-Leu (SEQ ID NO: 15), Pro-Val-Ala-Glu (SEQ ID NO: 16), Arg-Asn-Gly-Pro-Arg (SEQ ID NO: 17), Arg-Ala-Glu-Asp-Thr-Ala-Thr-Tyr-Tyr (SEQ ID NO: 18), Glu-Gly-Asp-Ser-Ala-Ala-Ile (SEQ ID NO: 19), Glu-Pro-Glu-Gly-Asp-Val-His-Gln (SEQ ID NO: 20), Glu-Asp-Glu-Val-Leu-Ala-Thr-Pro (SEQ ID NO: 21), and combinations thereof.

[0040] In a preferred embodiment, the protein hydrolyzate of the present invention is a protein hydrolyzate containing a peptide having at least an amino acid sequence selected from the group consisting of Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Glu-Phe-Asp-Glu-Lys-Pro-Ala-Asp (SEQ ID NO: 6), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Pro-Ile-Asn-Asp-Asn (SEQ ID NO: 14), and Pro-Thr-Asp-Leu (SEQ ID NO: 15).

[0041] The protein hydrolyzate of the present invention can be obtained or induced by enzymatic hydrolysis. In particular, the protein hydrolyzate is obtained by enzymatic hydrolysis of the blood of animals including, but not limited to, pigs, cows, and poultry (such as chickens, birds, turkeys, etc.). A preferred embodiment is that the animal blood is poultry blood. More preferably, the poultry blood is chicken blood.

[0042] The enzymatic hydrolysis for obtaining the protein hydrolyzate of the present invention is carried out using serine protease. Preferably, the serine protease is subtilisin.

[0043] The present invention also provides the use of the protein hydrolyzate defined above as a taste regulator for food preparations or pharmaceutical preparations.

[0044] A third aspect of the present invention relates to a taste regulating composition containing the above isolated peptide or protein hydrolyzate. The present invention also provides the use of the taste regulating composition in food preparations or pharmaceutical preparations. Accordingly, the present invention provides a food preparation or a pharmaceutical preparation containing the above isolated peptide, protein hydrolyzate, or taste regulating composition.

[0045] Furthermore, the present invention provides a method for modulating the taste of a food preparation, which comprises using, in an effective amount, the above-described isolated peptide, protein hydrolyzate, or taste-modulating composition containing an effective amount of the isolated peptide in a food preparation or a pharmaceutical preparation.

Examples

[0046] Hereinafter, the present invention will be described in more detail by the following non-limiting examples.

[0047] Example 1: Preparation and Identification of Peptides 1. Preparation of Peptides by Enzymatic Digestion All samples were prepared using chicken blood obtained from a slaughterhouse. First, the chicken blood was centrifuged at 4000 rpm for 8 minutes, and then digested with subtilisin enzyme (Protin SD-AY10) using 2M NaOH to adjust the initial pH to 9. The enzyme-digested sample was incubated at 55 °C and 120 rpm for 18 hours. Thereafter, the enzyme activity was stopped by heating in a water bath at 100 °C for 15 minutes and cooling to ambient temperature. The resulting solution was lyophilized and stored at -20 °C until use.

[0048] 2. Fractionation of Peptide Mixtures by Ultrafiltration Membranes 5 g of chicken blood powder was dissolved in 300 mL of deionized water in an ultrasonic bath. The resulting solution was transferred to an ultrafiltration device (8400 Millipore Amicon, EMD Millipore Corporation) equipped with a cut-off membrane of 1 kDa, 3 kDa, or 5 kDa (Ultacel® chemically regenerated cellulose (RC) filter with a diameter of 76 mm, EMD Millipore Corporation, Billerica, USA). The high molecular weight (HMW) and low molecular weight (LMW) fractions of each cut-off were washed with 50 mL of deionized water, lyophilized, and stored at -20 °C until further analysis. The yields of each cut-off membrane are summarized in Table 1, and it was found that the content of the compound was highest in the LMW fraction of each cut-off membrane.

[0049] [Table 1] TIFF2025518505000002.tif98169

[0050] To confirm which LMW fraction contains the most taste-active peptides and taste-modulating peptides, prior to sensory evaluation, the analysis of sodium and chloride contents in all LMW fractions of blood cells and plasma was performed by liquid chromatography using tandem mass spectrometry (LC-MS / MS). Quantitative analysis by ion chromatography showed that chicken blood contains little sodium and chloride, as none of the samples showed sodium and chloride exceeding 10% (Table 2). Plasma contains more sodium and chloride than blood cells, and the sodium and chloride contents of the 1 kDa LMW plasma were the highest among all samples.

[0051] [Table 2] TIFF2025518505000003.tif55169

[0052] The sensory evaluation was designed to clearly and comprehensively understand the taste characteristics of the samples. The HMW fraction was not used in the sensory experiment for safety reasons. The sensory samples were prepared by dissolving each LMW fraction of blood cells and plasma in the model broth to a final concentration of 10 g / L and adjusting the pH to 6 by adding 1% formic acid to water. The panelists were asked to evaluate the six taste qualities including umami, kokumi, sweetness, saltiness, sourness, and bitterness, as shown in Figures 1(a) and (b), on a scale of 0 (undetectable) to 5 (strongly detectable).

[0053] From the experimental results, it was found that the LMW of hydrolyzed chicken blood cells with a 3 kDa cut-off had significantly (P<0.01) taste activities and taste regulatory properties in terms of umami (3 kDa: 1.6), kokumi (3 kDa: 1.6), and bitterness (3 kDa: 2.0) compared to 1 kDa. The saltiness and sweetness were inferior. The enhancement of umami was comparable in both 1 kDa and 3 kDa cut-offs. Furthermore, the LMW fractions of 3 kDa and 5 kDa cut-offs were compared for both blood cells and plasma using the duo-trio test. The sensory tests were conducted in two series of AAB and BBA (the order was random). The panelists were asked to select a sample different from the other two samples within all sets. The duo-trio test designed to compare the 3 kDa hydrolyzed blood cell and the 3 kDa plasma LMW fractions was to select the sample with a more prominent taste compared to the other two samples. From the experimental results, no significant difference (P = 0.1) was observed, indicating that peptides larger than 3 kDa do not have significant taste activities and taste regulatory activities. The 3 kDa LMW plasma showed more prominent impressions of umami, kokumi, and saltiness compared to the LMW fraction of blood cells. Therefore, the 3 kDa cut-off LMW plasma fraction was selected for further research.

[0054] 3. Fractionation of 3 kDa LMW Plasma by Medium Pressure Liquid Chromatography (MPLC) 2 g of the 3 kDa LMW plasma powder was dissolved in 20 mL of deionized water, fractionated by reverse-phase (RP) MPLC (Buchi, Flawil, Switzerland) on a 150 × 40 mm polypropylene cartridge using a LiChroPrep RP18 bulk material (Merck, Darmstadt, Germany) with a particle size of 25 - 40 μm as the stationary phase and a gradient of 1% aqueous formic acid (solvent A) and methanol (solvent B) as the mobile phase. The fractionation was monitored using an evaporative light scattering detector (ELSD) (SEDERE, Alfortville Cedex, France). Chromatography (flow rate 40 mL / min) was carried out with 100% solvent A for 10 minutes, then the solvent B was increased to 30% within 15 minutes and further to 100% within 15 minutes, and then eluted with 100% solvent B for 15 minutes. Finally, seven fractions (F1 - F7) were collected, separated from the solvent in vacuo, and lyophilized twice before taste dilution analysis (TDA), comparative taste dilution analysis (cTDA), and chemical analysis (Figures 2(a), (b), (c)).

[0055] TDA was performed to find the most potent taste-active fraction, and cTDA was performed to find the taste-modulating fraction. In TDA, serial 1:2 dilutions of each fraction or sub-fraction were prepared with water and then presented to the sensory panelists in increasing concentration order. Each dilution was evaluated sensorially using the duo-trio test. The dilution at which the taste difference between the diluted fraction and two blanks (water) was just detected was defined as the taste dilution (TD) factor. The TD factors evaluated by five different panelists were averaged. The TD factors between individuals and between three individual sessions had a difference of less than one step in the dilution steps. Eight steps of dilution were performed (Oliver Frank, Jezussek, and Hofmann, 2003). For cTDA, the procedure was as described for TDA (Dunkel et al., 2007; Salger et al., 2019), except that the fractions were dissolved in model broth and the model broth was used as the blank. The comparative taste dilution (cTD) factor was defined by the dilution at which the taste difference between the diluted fraction and two blanks (model broth) could just be detected.

[0056] According to the results of TDA and cTDA, fraction F3 showed the highest taste activity and taste-modulating properties. In water, as shown in Fig. 2(a), fraction F2 showed the highest umami coefficient with a TD coefficient of 16, followed by fraction F3 (TD coefficient = 8) and fraction F4 (TD coefficient = 4). For saltiness, fractions F2 and F3 had the same TD coefficient of 4. Fraction F6 showed the highest bitterness and astringency (TD coefficient = 4). In the model broth, as shown in Fig. 2(b), fraction F3 showed the highest umami-enhancing activity (cTD factor = 16), followed by fraction F2 (cTD factor = 8) and fraction F4 (cTD factor = 4). The highest kokumi taste was found in fraction F4. Since none of the fractions showed bitterness in the model broth, it was suggested that fractions F2 - F5 were taste-enhancing fractions without an unpleasant characteristic taste (Fig. 2(c)). Since fractions F2, F3, and F4 had the highest TD and cTD coefficients, further analysis focused on these fractions.

[0057] 4. Fractionation of Fractions F2, F3, and F4 by High Performance Liquid Chromatography (HPLC) To identify the taste-active and taste-modulating peptides in fractions F2, F3, and F4, these fractions were further separated by preparative HPLC using a gradient of 1% aqueous formic acid and acetonitrile.

[0058] For the separation of fraction F2, 10 g of fraction F2 was dissolved in 1 L of deionized water and membrane-filtered (0.45 μm). 0.5 mL of this solution was injected into an HPLC apparatus (Jasco, Gros-Umstadt, Germany). The HPLC separation was carried out under the following conditions. Column: Luna HILIC (200A, 250 × 2.12 mm, 5 μm) Flow rate: 1.8 mL / min Detector: ELSD and UV (254 nm)

[0059] For the separation of fraction F3, 10 g of fraction F3 was dissolved in 1 L of deionized water and membrane-filtered (0.45 μm). 1 mL of this solution was injected into an HPLC apparatus (Jasco, Gros-Umstadt, Germany). Column: Luna PFP (2) (100A, 250×21.2 mm, 5 μm) Flow rate: 21 mL / min Detector: ELSD and UV (254 nm)

[0060] For the separation of fraction F4, 20 g of fraction F4 was dissolved in 1 L of deionized water and membrane filtered (0.45 μm). 1 mL of this solution was injected into an HPLC apparatus (Jasco, Gros-Umstadt, Germany). Column: Nucleodur C18 Pyramid (100A, 250×21.2 mm, 5 μm) Flow rate: 20 mL / min Detector: ELSD and UV (254 nm)

[0061] The sub-fractions of F2, F3, and F4 were analyzed using a liquid chromatography time-of-flight mass spectrometer (LC-TOF-MS) with a C8 column, and peptides were identified using PEAKS software.

[0062] From the experimental results, since fraction F2 contained known taste-active and taste-modulating molecules (e.g., cations, amino acids, and glutamyl dipeptides) at high concentrations, fractions F3 and F4 were subjected to large-scale separation in order to identify novel taste-active and taste-modulating peptides.

[0063] Example 2: Identification and Evaluation of Taste Activity and Taste Modulating Effects of Peptides 1. Identification of Peptides After analyzing the HPLC sub-fractions of MPLC fractions F2, F3, and F4, potential taste-active and taste-modulating peptides present in fractions F3 and F4 were identified using a combination of proteomics and sensomics techniques (so-called sensoproteomics approach) from the method described in Sebald et al., J. Agric. Food Chem. 2018, 66, 11092-11104. Both targeted proteomics and untargeted proteomics are applied to improve the productivity of the present invention.

[0064] Using the target proteomics method, all the peptides theoretically obtained from chicken serum albumin and corticosteroid-binding globulin were examined, and 767 peptides were detected. According to the non-target proteomics method using Maxquant and PEAKS software, peptides in the LC-TOF-MS data of the sub-fractions of MPLC fractions F2, F3 and F4 were identified. Among the peptides identified in fractions F3 and F4 using de novo sequencing and database search of PEAKS, all those with an ALC (average local confidence) of 80 or more and an N-terminus of E, P or R were selected. Using Maxquant, first the taste-active fraction and taste-modulating fraction were examined, and then the identified peptides were selected according to the N-terminus E, P or R and peak area. Eleven peptides were obtained from fraction F3 and 44 peptides were obtained from fraction F4, which were further confirmed by MS 2 data.

[0065] After identifying the peptides, several filtration steps were performed, and then peptide candidates were selected for verification. The final peptide candidates identified in MPLC fractions F3 and F4 are shown in Table 3 below.

[0066] [Table 3] TIFF2025518505000004.tif62169

[0067] TIFF2025518505000005.tif145169

[0068] TIFF2025518505000006.tif149169

[0069] TIFF2025518505000007.tif134169

[0070] TIFF2025518505000008.tif52169

[0071] Twenty-one candidate peptides were synthesized, and their purities were quantified 1It was evaluated to be in the range of 62% to 100% by 1H-NMR. The presence of peptides in the sample was confirmed by LC-MS / MS using Skyline. Comparing the transitions of MRM (Multiple Reaction Monitoring) detected in MPLC fraction F3 or F4 with synthetic peptides, it was possible to confirm the presence of candidate peptides in the plasma of chickens hydrolyzed according to the present invention.

[0072] 2. Determination of Taste Thresholds of Peptides The threshold concentrations of the newly identified 21 peptides were measured in water for the inherent taste and in model broth (adjusted to pH 6.0 with 1% formic acid) for the taste regulatory properties, respectively (Table 4). The peptides were dissolved and serially diluted 1:2, and presented to the panelists using the duo-trio test while increasing the concentration as reported in the literature. The geometric mean of the last concentration and the second last concentration was calculated and used as the individual recognition threshold. The threshold of the sensory panel was approximated by averaging the individual thresholds in two independent measurements.

Number

Number

[0073] [Table 4] TIFF2025518505000011.tif121169

[0074] TIFF2025518505000012.tif174169

[0075] TIFF2025518505000013.tif132169

[0076] From the results, the taste threshold concentration was in the range of 100 - 1024 μmol / L in water. Among the 21 peptides, 7 did not show any specific taste up to a concentration of 2 mmol / L (i.e., the peptides having SEQ ID NO: 1, 6, 8, 14, 15, 17, and 19).

[0077] Furthermore, all 21 peptides showed taste-modulating properties in the model broth in the range of 26 μmol / L - 351 μmol / L. The peptide having SEQ ID NO: 14 had the lowest umami and kokumi taste thresholds at 26 μmol / L. The peptide having SEQ ID NO: 8 had the property of enhancing kokumi and salty tastes, and the highest threshold was 351 μmol / L. The peptides having SEQ ID NO: 3, 7, and 20 had the properties of modulating kokumi, umami, and salty tastes at thresholds of 236, 95, and 61 μmol / L, respectively. The peptides having SEQ ID NO: 12, 17, 18, 19, and 20 showed the property of enhancing kokumi and umami tastes at thresholds of 137, 101, 247, 212, and 61 μmol / L, respectively.

[0078] Furthermore, peptides having SEQ ID NO: 1, 3, 6, 8, 11, 13, 14, 16, and 21, which showed the property of modulating salty taste, were selected, and an isointensity test of saltiness was conducted to evaluate the property of enhancing or reducing salty taste.

[0079] 3. Isointensity Tests of Saltiness of Peptides To examine the effect of the peptides on the perceived salty taste, an isointensity test of saltiness was conducted according to the literature. In the isointensity test of saltiness, the peptide (1 mmol / L) was mixed into a model broth containing 50 mM NaCl, and the panelists were asked to evaluate the intensity of the salty taste compared with the following NaCl concentrations: 30, 40, 50, 60, 70, and 80 mmol / L. The results were as shown in Table 5.

[0080] [Table 5] TIFF2025518505000014.tif128169

[0081] TIFF2025518505000015.tif57169

[0082] The perceived isointensity of saltiness for all peptides was in the range of 42.9 - 66.7 mmol / L of NaCl concentration in the model broth. Peptides with SEQ ID NO: 6, 8, 11, 13, 14, 16, and 21 showed saltiness enhancing properties, while peptides with SEQ ID NO: 1, 3, and 13 showed saltiness reducing properties. The strongest saltiness enhancing effect was observed in the peptide with SEQ ID NO: 8, which had a 33% saltiness enhancement (isointensity of saltiness perceived at 50 - 66.7 mmol / L), and the lowest saltiness enhancing effect was observed in the peptide with SEQ ID NO: 21, which had a 20% saltiness enhancement (isointensity of saltiness perceived at 50 - 59.8 mmol / L). Furthermore, for the peptides with SEQ ID NO: 1, 3, and 13, saltiness reducing effects of 14% and 13% were observed, respectively.

[0083] 4. Identification of Major Taste Active and Taste Modulating Peptides To identify peptides that play important roles in taste activity and taste modulation effects, not only the newly identified 21 peptides but also basic seasonings known in the literature (e.g., amino acids, γ-glutamyl peptides, cations, anions, etc.) were quantified in hydrolyzed chicken plasma samples. Focusing on the newly identified 21 peptides, Table 6 shows the taste quality, taste threshold, dose threshold (DoT) coefficient, and peptide concentration in hydrolyzed chicken plasma samples. The taste threshold determined for the newly identified peptides (listed in Table 6) was used in the calculation of the DoT coefficient, which is defined as the ratio of the concentration of a substance to the corresponding taste threshold concentration. Since substances with a DoT coefficient ≧ 1 are particularly likely to directly contribute to the perceived taste, the DoT coefficient enables the initial estimation of the contribution to taste.

[0084] [Table 6] TIFF2025518505000016.tif212169

[0085] TIFF2025518505000017.tif86169

[0086] From the results, five peptides (SEQ ID NOs: 3, 6, 11, 14, and 15) having a DoT coefficient ≧ 1 played important roles in the taste activity and taste regulatory effects in plasma samples.

[0087] Example 3: Taste Modulating Effects of Peptides Identified from Foods To examine the taste regulatory performance of peptides identified from foods, further sensory experiments were conducted using commercially available chicken soup and model broth. In the sensory experiments, peptides such as Pro-Val-Leu-Lys (SEQ ID NO: 1), Pro-Leu-Pro-Arg (SEQ ID NO: 2), Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Pro-Gln-Pro-Glu-Arg (SEQ ID NO: 5), and Glu-Asp-Ala-Pro-Val-Leu-His (SEQ ID NO: 7) were mixed into commercially available chicken soup (Netto Marken-Discount AG&CO.KG, Maxhutte-Haidhof) and model broth. In the experiment, 3 g of commercially available chicken soup powder was dissolved in 150 mL of boiling water. The peptide (2 mmol / L each) was mixed into 20 mL of this solution. Then, the taste threshold was determined by a duo-trio test as described above (using chicken soup as a reference). Table 7 shows the sensory evaluation of the identified peptides in commercially available chicken soup and model broth.

[0088] [Table 7] TIFF2025518505000018.tif81169

[0089] From the results, it was found that the peptides having SEQ ID NOs: 1, 3, and 7 can affect the taste characteristics of commercially available chicken soup at about 1500 μmol / L. In the case of the peptide having SEQ ID NO: 5, the mixed samples could not be distinguished (the maximum concentration tested was 2 mmol / L). The peptide having SEQ ID NO: 2 could be perceived at a concentration of 418 μmol / L in commercially available chicken soup, but the threshold was 4 times lower (112 μmol / L) in the model broth. Based on these experimental results, it can be concluded that the peptides having specific amino acid sequences according to the present invention can enhance the taste profile of commercially available foods, although higher concentrations are required compared to the model broth.

[0090] Furthermore, additional commercially available foods having different matrices, such as chicken soup (Netto Marken-Discount AG&CO.KG, Maxhutte-Haidhof), vegetable soup (Hugli Nahrungsmittel GmbH, Radolfzell), and pepper sauce (Unilever Deutschland, Hamburg) purchased from local supermarkets, were used to evaluate the taste-modulating effects of the peptides newly identified according to the present invention, for example, Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Glu-Phe-Asp-Glu-Lys-Pro-Ala-Asp (SEQ ID NO: 6), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Pro-Ile-Asn-Asp-Asn (SEQ ID NO: 14), and Pro-Thr-Asp-Leu (SEQ ID NO: 15). Food samples were prepared according to the detailed descriptions provided for each product. For chicken soup and vegetable soup, 3 g of the sample was dissolved in 150 mL of boiling water and cooled to room temperature in an ice bath. For pepper sauce, 4 g was dissolved in 150 mL of boiling water. Then, 200 μmol / L of the peptide was mixed into 20 mL of the soup or sauce. The 3-AFC test was designed to test the taste characteristics of the peptides in each commercially available product. Two of the samples contained only 20 mL of the commercially available product, and one sample had the peptide added to 20 mL of the commercially available product. Panelists were asked to select the different samples. Table 8 shows the results of the sensory tests for three commercially available foods mixed with five exemplary peptides.

[0091] [Table 8] TIFF2025518505000019.tif39169

[0092] The results clearly show that the peptides newly identified according to the present invention can be used as taste modulators in edible products, particularly food preparations or even pharmaceutical preparations.

[0093] Example 4: Effects of Enzyme Types on Peptide Sequences To examine the effect of the type of enzyme on the amino acid sequences of peptides present in a hydrolyzed plasma sample, chicken plasma was digested with various enzymes, such as flavase and thermolysin.

[0094] For digestion with flavase, 0.0504 g of chicken plasma was extracted three times with methanol, and the sample was dried under nitrogen. The sample was redissolved in 1 M TEAB, and urea was added to a final concentration of 8 M. Next, 100 mM DTT was added, and the mixture was maintained at 56 °C for 45 minutes. Subsequently, 0.55 M IAA was added, and the mixture was incubated at room temperature in the dark for 1 hour. Next, the sample was digested with flavase at 37 °C for 15 or 30 minutes. The digestion was stopped by adding concentrated formic acid, and the peptide mixture was purified by SPE on a C18 cartridge.

[0095] For digestion with thermolysin, the plasma sample was prepared by pre-adjusting the pH to 8 and then incubated with thermolysin at 55 °C for 18 hours. The reaction was stopped by heating in a 100 °C water bath for 15 minutes. The solution was further dried before analysis.

[0096] Peptide identification was performed by de novo sequencing of LC-TOF-MS data using PEAKS to screen for 21 peptides (SEQ ID NOs: 1-21, Table 3) in the hydrolyzed chicken plasma sample as described above. Due to the high activity of flavase, chicken plasma was digested with flavase for 15 or 30 minutes.

[0097] Table 9 shows exemplary peptides identified in chicken plasma after digestion with flavase, and Table 10 shows exemplary peptides identified in chicken plasma after digestion with thermolysin.

[0098] [Table 9] TIFF2025518505000020.tif116169

[0099] TIFF2025518505000021.tif153169

[0100] TIFF2025518505000022.tif27169

[0101] In the case of digestion by flavorzyme, it was found that the length of the peptides identified by 15-minute digestion was longer compared to 30-minute digestion. The longer the time, the more thoroughly the digestion proceeds, resulting in an increase in the number of shorter peptides.

[0102] [Table 10] TIFF2025518505000023.tif43169

[0103] Furthermore, for 21 peptides (SEQ ID NOs: 1 to 21, Table 3), which are new peptides identified by the present invention, targeted LC-MS / MS analysis was performed on the digests by flavorzyme and thermolysin. As a result, it was revealed that none of these peptides were present because the behaviors of the enzymes were different.

Claims

1. Pro-Val-Leu-Lys (SEQ ID NO: 1), Pro-Leu-Pro-Arg (SEQ ID NO: 2), Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Pro-Glu-Arg-Asn-Glu (SEQ ID NO: 4), Pro-Gln-Pro-Glu-Arg (SEQ ID NO: 5), Glu-Phe-Asp-Glu-Lys-Pro-Ala-Asp (SEQ ID NO: 6), Glu-Asp-Ala-Pro-Val-Leu-His (SEQ ID NO: 7), Glu-Phe-Asp-Ala-Arg-Pro-Thr (SEQ ID NO: 8), Glu-Glu-Thr-Pro-Ser-His-Asp (SEQ ID NO: 9), Glu-Glu-Val-Glu-Glu-Glu-Glu-Val-Glu (SEQ ID NO: 10), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Glu-Asn-Ala-Pro-Asp-Gln-Lys (SEQ ID NO: 12), Glu-Asp-Met-Ala-Pro (SEQ ID NO: 13), Pro-Ile-Asn-Asp-Asn (SEQ ID NO: 14), Pro-Thr-Asp-Leu (SEQ ID NO: 15), Pro-Val-Ala-Glu (SEQ ID NO: 16), Arg-Asn-Gly-Pro-Arg (SEQ ID NO: 17), Arg-Ala-Glu-Asp-Thr-Ala-Thr-Tyr-Tyr (SEQ ID NO: 18), Glu-Gly-Asp-Ser-Ala-Ala-Ile (SEQ ID NO: 19), Glu-Pro-Glu-Gly-Asp-Val-His-Gln (SEQ ID NO: 20), and An isolated peptide having an amino acid sequence selected from the group consisting of Glu-Asp-Glu-Val-Leu-Ala-Thr-Pro (SEQ ID NO: 21). **Claim 2** The isolated peptide according to claim 1, which imparts at least one of a saltiness-reducing effect, a saltiness-enhancing effect, an umami effect, and a kokumi effect. **Claim 3** Pro-Val-Leu-Lys (SEQ ID NO: 1), Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), and The isolated peptide according to claim 1 or 2, which has an amino acid sequence selected from the group consisting of Glu-Asp-Met-Ala-Pro (SEQ ID NO: 13) and imparts a saltiness-reducing effect. **Claim 4** Glu-Phe-Asp-Glu-Lys-Pro-Ala-Asp (SEQ ID NO: 6), Glu-Phe-Asp-Ala-Arg-Pro-Thr (SEQ ID NO: 8), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Pro-Ile-Asn-Asp-Asn (SEQ ID NO: 14), Pro-Val-Ala-Glu (SEQ ID NO: 16), and The isolated peptide according to claim 1 or 2, which has an amino acid sequence selected from the group consisting of Glu-Asp-Glu-Val-Leu-Ala-Thr-Pro (SEQ ID NO: 21) and imparts a saltiness-enhancing effect. **Claim 5** Pro-Val-Leu-Lys (SEQ ID NO: 1), Pro-Leu-Pro-Arg (SEQ ID NO: 2), Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Pro-Glu-Arg-Asn-Glu (SEQ ID NO: 4), Pro-Gln-Pro-Glu-Arg (SEQ ID NO: 5), Glu-Phe-Asp-Glu-Lys-Pro-Ala-Asp (SEQ ID NO: 6), Glu-Asp-Ala-Pro-Val-Leu-His (SEQ ID NO: 7), Glu-Glu-Thr-Pro-Ser-His-Asp (SEQ ID NO: 9), Glu-Glu-Val-Glu-Glu-Glu-Glu-Val-Glu (SEQ ID NO: 10), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Glu-Asn-Ala-Pro-Asp-Gln-Lys (SEQ ID NO: 12), Pro-Ile-Asn-Asp-Asn (SEQ ID NO: 14), Arg-Asn-Gly-Pro-Arg (SEQ ID NO: 17), Arg-Ala-Glu-Asp-Thr-Ala-Thr-Tyr-Tyr (SEQ ID NO: 18), Glu-Gly-Asp-Ser-Ala-Ala-Ile (SEQ ID NO: 19), Glu-Pro-Glu-Gly-Asp-Val-His-Gln (SEQ ID NO: 20), and Glu-Asp-Glu-Val-Leu-Ala-Thr-Pro (SEQ ID NO: 21), and has an amino acid sequence selected from the group consisting of, imparting umami effect, the isolated peptide according to claim 1 or 2.

6. Pro-Val-Leu-Lys (SEQ ID NO: 1), Pro-Leu-Pro-Arg (SEQ ID NO: 2), Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Pro-Glu-Arg-Asn-Glu (SEQ ID NO: 4), Pro-Gln-Pro-Glu-Arg (SEQ ID NO: 5), Glu-Asp-Ala-Pro-Val-Leu-His (SEQ ID NO: 7), Glu-Phe-Asp-Ala-Arg-Pro-Thr (SEQ ID NO: 8), Glu-Glu-Thr-Pro-Ser-His-Asp (SEQ ID NO: 9), Glu-Glu-Val-Glu-Glu-Glu-Glu-Val-Glu (SEQ ID NO: 10), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Glu-Asn-Ala-Pro-Asp-Gln-Lys (SEQ ID NO: 12), Glu-Asp-Met-Ala-Pro (SEQ ID NO: 13), Pro-Thr-Asp-Leu (SEQ ID NO: 15), Pro-Val-Ala-Glu (SEQ ID NO: 16), Arg-Asn-Gly-Pro-Arg (SEQ ID NO: 17), Arg-Ala-Glu-Asp-Thr-Ala-Thr-Tyr-Tyr (SEQ ID NO: 18), Glu-Gly-Asp-Ser-Ala-Ala-Ile (SEQ ID NO: 19), Glu-Pro-Glu-Gly-Asp-Val-His-Gln (SEQ ID NO: 20), and Glu-Asp-Glu-Val-Leu-Ala-Thr-Pro (SEQ ID NO: 21), and having an amino acid sequence selected from the group consisting of, imparting kokumi effect, the isolated peptide according to claim 1 or 2.

7. An isolated peptide according to any one of claims 1 to 6, obtained by enzymatic hydrolysis or fermentation.

8. An isolated peptide according to claim 7, obtained by enzymatic hydrolysis of animal blood.

9. The isolated peptide according to claim 8, wherein the animal blood is poultry blood.

10. The isolated peptide according to claim 8, wherein the enzymatic hydrolysis is carried out using a serine protease.

11. The isolated peptide according to claim 10, wherein the serine protease is subtilisin.

12. Use of the isolated peptide according to any one of claims 1 to 11 as a taste modifier in a food preparation or a pharmaceutical preparation.

13. Pro-Val-Leu-Lys (SEQ ID NO: 1), Pro-Leu-Pro-Arg (SEQ ID NO: 2), Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Pro-Glu-Arg-Asn-Glu (SEQ ID NO: 4), Pro-Gln-Pro-Glu-Arg (SEQ ID NO: 5), Glu-Phe-Asp-Glu-Lys-Pro-Ala-Asp (SEQ ID NO: 6), Glu-Asp-Ala-Pro-Val-Leu-His (SEQ ID NO: 7), Glu-Phe-Asp-Ala-Arg-Pro-Thr (SEQ ID NO: 8), Glu-Glu-Thr-Pro-Ser-His-Asp (SEQ ID NO: 9), Glu-Glu-Val-Glu-Glu-Glu-Glu-Val-Glu (SEQ ID NO: 10), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Glu-Asn-Ala-Pro-Asp-Gln-Lys (SEQ ID NO: 12), Glu-Asp-Met-Ala-Pro (SEQ ID NO: 13), Pro-Ile-Asn-Asp-Asn (SEQ ID NO: 14), Pro-Thr-Asp-Leu (SEQ ID NO: 15), Pro-Val-Ala-Glu (SEQ ID NO: 16), Arg-Asn-Gly-Pro-Arg (SEQ ID NO: 17), Arg-Ala-Glu-Asp-Thr-Ala-Thr-Tyr-Tyr (SEQ ID NO: 18), Glu-Gly-Asp-Ser-Ala-Ala-Ile (SEQ ID NO: 19), Glu-Pro-Glu-Gly-Asp-Val-His-Gln (SEQ ID NO: 20), Glu-Asp-Glu-Val-Leu-Ala-Thr-Pro (SEQ ID NO: 21), and A protein hydrolyzate comprising at least one peptide having an amino acid sequence selected from the group consisting of these combinations.

14. Glu-Ala-Glu-Phe-Asp (SEQ ID NO: 3), Glu-Phe-Asp-Glu-Lys-Pro-Ala-Asp (SEQ ID NO: 6), Glu-Phe-Asp-Glu-Lys-Ala-Pro-Asp (SEQ ID NO: 11), Pro-Ile-Asn-Asp-Asn (SEQ ID NO: 14), and The protein hydrolyzate according to claim 13, comprising at least a peptide having an amino acid sequence selected from the group consisting of Pro-Thr-Asp-Leu (SEQ ID NO: 15).

15. The protein hydrolyzate according to claim 13 or 14, obtained by enzymatic hydrolysis.

16. The protein hydrolyzate according to claim 15, obtained by enzymatic hydrolysis of animal blood.

17. The protein hydrolyzate according to claim 16, wherein the animal blood is poultry blood.

18. The protein hydrolyzate according to claim 16, wherein the enzymatic hydrolysis is performed using a serine protease.

19. The protein hydrolyzate according to claim 18, wherein the serine protease is subtilisin.

20. Use of the protein hydrolyzate according to any one of claims 13 to 19 as a taste modifier in a food preparation or a pharmaceutical preparation.

21. A taste modulating composition comprising the isolated peptide according to any one of claims 1 to 11.

22. A taste-modulating composition comprising the protein hydrolyzate according to any one of claims 13 to 19.

23. Use of the taste-modulating composition according to claim 21 or 22 in a food preparation or a pharmaceutical preparation.

24. A food preparation comprising the isolated peptide according to any one of claims 1 to 11.

25. A food preparation comprising the protein hydrolyzate according to any one of claims 13 to 19.

26. A food preparation comprising the taste-modulating composition according to claim 21 or 22.

27. A pharmaceutical preparation comprising the isolated peptide according to any one of claims 1 to 11.

28. A pharmaceutical preparation comprising the protein hydrolyzate according to any one of claims 13 to 19.

29. A pharmaceutical preparation comprising the taste-modulating composition according to claim 21 or 22.

30. A method for modulating the taste of a food preparation, comprising applying an effective amount of the isolated peptide according to any one of claims 1 to 11 to the food preparation.

31. A method for modulating the taste of a food preparation, comprising applying to the food preparation a protein hydrolyzate according to any one of claims 13 to 19, which contains an effective amount of the isolated peptide.

32. A method for modulating the taste of a food preparation, comprising applying to the food preparation a taste-modulating composition according to claim 21 or 22, which contains an effective amount of the isolated peptide.

33. A method for modulating the taste of a pharmaceutical preparation, comprising applying an effective amount of the isolated peptide according to any one of claims 1 to 11 to the pharmaceutical preparation.

34. A method for modulating the taste of a pharmaceutical preparation, comprising applying a protein hydrolysate according to any one of claims 13 to 19, which contains an effective amount of an isolated peptide, to the pharmaceutical preparation. **Claim 35** A method for modulating the taste of a pharmaceutical preparation, comprising applying a taste-modulating composition according to claim 21 or 22, which contains an effective amount of an isolated peptide, to the pharmaceutical preparation.

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