Novel separating peptide, protein hydrolyze comprising same, and use thereof

Abstract

The present invention relates to novel isolated peptides, protein hydrolysates comprising said isolated peptides for use as taste modifiers in food and pharmaceutical formulations. The present invention relates to an isolated peptide having an amino acid sequence selected from the group listed herein, to protein hydrolysates comprising said isolated peptide, and to their use as taste modulators in food and pharmaceutical formulations.

Description

Technical Field

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

[0002] Livestock blood is a byproduct obtained from industrial slaughtering processes. Many countries around the world produce and waste a large amount of livestock blood, such as chicken blood, every year, despite the fact that these bloods contain valuable proteins and bioavailable nutrients. Therefore, the use of these bloods is beneficial to both the economy and the environment. So far, the use of livestock blood has been limited in terms of cost-effectiveness and feeding goals, because the use of whole blood and protein in the food industry is restricted by their unpleasant taste or color. In order to address these problems, some research has been conducted to recycle byproducts and waste materials obtained from poultry processing and industry. For example, chicken blood can be used to make blood sausage, blood tofu and blood pudding. Plasma can be used as a protein supplement, an excellent source of trace elements, or as a substitute for protein in the baking industry due to its strong foaming ability (Jayathilakan et al., J Food Sci Technol, 2012, 49 (3), 378-293).

[0003] To improve the utilization of chicken blood, it is suggested to generate taste-active peptides and taste-modulating peptides. However, the existing knowledge of the composition of chicken blood (e.g., which proteins are present or the exact concentrations of several components) is limited. In addition, although many studies have been conducted to identify taste-active peptides and taste-modulating peptides in various foods, there are no studies related to the taste of chicken blood. Due to the limited knowledge and the complex matrix of chicken blood, it is challenging to identify taste-active peptides and taste-modulating peptides from livestock and poultry blood (e.g., chicken blood).

[0004] CN108047313A discloses a chicken blood cell antioxidant peptide and a fermentation preparation method thereof.

[0005] The amino acid sequence of the antioxidant peptide is Thr-Ser-Phe-Gly-Asp-Ala-Val-Lys-Asn-Leu-Asp-Asn-Ile-Lys (SEQ ID NO: 1). The antioxidant peptide has high free radical scavenging activity, high scavenging effect on DPPH free radicals, and strong reducing ability. The method includes the steps of culture activation, blood cell culture medium preparation, inoculation fermentation, separation and purification.

[0006] CN108048518A discloses a chicken blood cell antioxidant peptide and an enzymatic preparation method thereof.

[0007] The amino acid sequence of the antioxidant peptide is 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, high scavenging effect on DPPH free radicals and superoxide anion free radicals, and strong reducing ability. The method includes the steps of blood cell preparation, blood cell lysis, protein hydrolysis and ultrafiltration.

[0008] CN101843289A discloses a method for producing blood polypeptide protein powder using livestock and poultry blood. According to the method, the macromolecular albumin in livestock and poultry blood that is difficult to absorb and utilize is digested into small molecular albumin, polypeptides and free amino acids to improve the palatability and nutritional value of the dried blood for feeding.

[0009] Although the above-mentioned products and methods utilizing livestock and poultry blood have been developed, there is no disclosure of utilizing livestock and poultry blood, especially livestock and poultry blood-derived peptides having a specific amino acid sequence, as flavor active agents and flavor modifiers. Summary of the invention

[0010] The present invention has been invented in view of the above-mentioned problems.

[0011] Therefore, the object of the present invention is to utilize animal blood, in particular livestock blood as industrial waste from the slaughtering process, to obtain new taste active peptides and mixtures thereof for use as taste modifiers in food and pharmaceutical preparations.

[0012] Another object of the present invention is to provide animal blood-derived peptides, especially livestock and poultry blood-derived peptides, having specific amino acid sequences. The animal blood-derived peptides of the present invention can be used as taste activators to impart taste-modulating effects, such as salt reduction, salt increase, umami and kokumi effects.

[0013] Another object of the present invention is to provide a protein hydrolyzate and a taste-adjusting composition containing the animal blood-derived peptide.

[0014] Another object of the present invention is to provide a method for adjusting the taste of foods and pharmaceutical preparations by using the animal blood-derived peptide, protein hydrolyzate and taste adjusting composition.

[0015] In one aspect, the 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); 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).

[0016] 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: 12); 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.

[0017] In another aspect, the present invention relates to a taste modifying composition comprising said isolated peptide or protein hydrolysate as defined above.

[0018] In another aspect, the present invention relates to a food or pharmaceutical preparation comprising the isolated peptide, protein hydrolysate or taste modifying composition defined above.

[0019] Furthermore, the present invention also relates to a method for adjusting the taste of a food or a pharmaceutical preparation, comprising applying an effective amount of the isolated peptide, protein hydrolyzate or taste adjusting composition defined hereinabove to the food or the pharmaceutical preparation. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] The characteristics of the invention will be more clearly described on reading the following description of an exemplary embodiment, which is intended to be illustrative and not restrictive.

[0021] Figure 1 Shown are the sensory profiles of: (a) the low molecular weight (LMW) fraction of hydrolyzed chicken blood cells in model broth (MB) at a concentration of 10 g / L; and (b) the low molecular weight fraction of hydrolyzed chicken plasma in model broth at a concentration of 10 g / L.

[0022] Figure 2 Shown are RP 18 MPLC-ELSD chromatograms of MPLC fractions F2-F7 in water (a), model broth (MB) (b), and in water or model broth (c) and taste dilution comparison analysis. DETAILED DESCRIPTION

[0023] Unless otherwise stated, any aspect shown herein should also include application to other aspects of the invention.

[0024] Unless defined otherwise, technical and scientific terms used herein have the same definitions as understood by those skilled in the art.

[0025] In the present invention, the term "about" is used to indicate that any numerical value shown or presented herein may vary or deviate. Such variations or deviations may be caused by errors in the equipment or method used to determine the numerical value.

[0026] The terms "consisting of" and its variations, "comprising" and its variations, "having", "including" and its variations are open-ended verbs. For example, any method "consisting of", "comprising", "having" or "including" one or more components or steps is not limited to the one or more components or steps but also includes components or steps not mentioned.

[0027] Unless otherwise stated, "a", "an", "the", when referring to the singular form, are intended to include the plural form of the noun as well.

[0028] The term "taste adjustment" as used herein is synonymous with the terms "taste improvement", "taste adjustment", "taste reduction", "taste enhancement" and "taste change" and can be used interchangeably. According to the present invention, the terms "taste adjuster" and "taste adjustment composition" refer to agents and compositions that can impart one or more tastes (e.g., salty, sweet, kokumi, umami, astringent, bitter, sour, etc.) to food or pharmaceutical preparations and can improve the taste of food or pharmaceutical preparations as needed. In addition, taste adjustment can also be achieved by reducing or enhancing one or more of the tastes.

[0029] The term "food preparation" as used herein refers to all food and beverage products, compositions and preparations edible to humans and animals, including, for example, meals, beverages, dietary supplements, snacks, desserts, candies. Similarly, the term "pharmaceutical preparation" as used herein refers to all edible pharmaceutical products, compositions and preparations. Food and pharmaceutical preparations can be in solid form, such as powders, tablets, granules, beads, capsules, etc., or in semi-solid, gel and liquid forms.

[0030] Any tools, equipment, methods, materials or chemicals mentioned herein, unless otherwise specified, are tools, equipment, methods, materials or chemicals commonly used or practiced by those skilled in the art.

[0031] All disclosed chemicals, compounds, materials, components and / or methods and the claims of the present invention are intended to cover all aspects of the present invention obtained by any operation, practice, improvement or change of various factors without conducting experiments that are obviously different from the present invention, and are intended to obtain objects that have similar properties, functions and effects to the various aspects of the present invention according to the opinions of ordinary technicians in the field, although not specifically specified in the claims. Therefore, objects that are equivalent or similar to the various aspects of the present invention, including any minor improvements or changes that are obvious to ordinary technicians in the field, should also be considered to be within the spirit, scope and concept of the present invention.

[0032] Hereinafter, the present invention will be described in more detail.

[0033] According to a first aspect of the invention, the isolated peptide 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-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); 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: 12); 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 IDNO: 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).

[0034] The isolated peptide of the present invention can impart at least one of the following effects: salt reduction, salt increase, umami and kokumi effects.

[0035] In a preferred aspect, the isolated peptide that confers a salt-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).

[0036] In another preferred aspect, the isolated peptide that confers a salt-increasing 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).

[0037] In another preferred aspect, the isolated peptide that imparts 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).

[0038] In another preferred aspect, the isolated peptide that imparts a kokumi 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-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).

[0039] The isolated peptides of the present invention are obtained or derived from enzymatic hydrolysis or fermentation. Specifically, the isolated peptides are obtained or derived from enzymatic hydrolysis of animal blood (including but not limited to pigs, cattle and poultry (such as chickens, birds, turkeys, etc.)). In a preferred aspect, the animal blood useful for the present invention is poultry blood, more preferably chicken blood.

[0040] In an exemplary embodiment, the isolated peptide of the present invention is obtained by enzymatic hydrolysis using a serine protease. Preferably, the serine protease is subtilisin.

[0041] The present invention also provides the use of the isolated peptide as defined above as a taste modifier in food or pharmaceutical preparations.

[0042] A second aspect of 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: 10); 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: 12); 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.

[0043] In a preferred embodiment, the protein hydrolyzate of the present invention comprises peptides having at least the following amino acid sequences: 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).

[0044] The protein hydrolysate of the present invention can be obtained or derived by enzymatic hydrolysis. In particular, the protein hydrolysate is obtained by enzymatic hydrolysis of animal blood (including but not limited to pigs, cattle and poultry (such as chickens, birds, turkeys, etc.)). In a preferred aspect, the animal blood is poultry blood. More preferably, the poultry blood is chicken blood.

[0045] The protein hydrolysate of the present invention is obtained by enzymatic hydrolysis using a serine protease. Preferably, the serine protease is subtilisin.

[0046] The present invention also provides the use of a protein hydrolysate as defined herein before as a taste modifier for food or pharmaceutical preparations.

[0047] A third aspect of the present invention relates to a taste-adjusting composition comprising the isolated peptide or protein hydrolysate as defined above. The present invention also provides the use of the taste-adjusting composition in a food or pharmaceutical preparation. Therefore, the present invention provides a food or pharmaceutical preparation comprising the isolated peptide, protein hydrolysate or taste-adjusting composition as defined above.

[0048] In addition, the present invention provides a method for adjusting the taste of a food preparation, comprising applying an effective amount of the isolated peptide, protein hydrolyzate or a taste adjusting composition comprising an effective amount of the isolated peptide to a food or a pharmaceutical preparation.

[0049] Example

[0050] The present invention will be described in more detail below by means of the following non-limiting examples.

[0051] Example 1: Preparation and identification of peptides

[0052] 1. Preparation of Peptides by Enzymatic Digestion

[0053] All samples were prepared from slaughterhouse chicken blood. First, the chicken blood was centrifuged at 4000 rpm for 8 minutes and then digested with subtilisin (Protin SD-AY 10) and the initial pH was adjusted to 9 with 2M NaOH. The enzyme-digested samples were incubated at 55°C at 120 rpm for 18 hours. Afterwards, the enzymes were inactivated by heating in a 100°C water bath for 15 minutes and cooling to ambient temperature. The resulting solution was freeze-dried and stored at -20°C until further use.

[0054] 2. Fractionation of the Peptide Mixture by Ultrafiltration

[0055] 5 g of chicken blood plasma powder was dissolved in 300 ml of deionized water in an ultrasonic bath. The resulting solution was transferred to an ultrafiltration system (8400 Millipore Amicon, EMD Millipore) equipped with a 1, 3 or 5 kDa cut-off membrane (76 mm diameter) and a 100 μm ultrafiltration membrane (100 μm). Chemically regenerated cellulose (RC) filters, EMD Milliporecorporation, Billerica, USA). Each retained high molecular weight (HMW) and low molecular weight (LMW) fraction was washed with 50 mL of deionized water, freeze-dried and stored at -20 °C until further analysis. The yields of the different membrane cutoffs are summarized in Table 1, and the compounds with the highest content in each low molecular weight cutoff fraction are emphasized.

[0056] Table 1

[0057]

[0058] To determine which low molecular weight fraction contained the highest content of taste active and taste modulating peptides, all low molecular weight fractions in blood cells and plasma were analyzed for sodium and chloride content prior to sensory evaluation and analyzed using liquid chromatography-tandem mass spectrometry (LC-MS / MS). Quantitative analysis by ion chromatography showed that the sodium and chloride content in chicken blood was not high, as none of the samples showed more than 10% sodium and chloride (Table 2). The sodium and chloride content of plasma samples was higher than that of blood cells, and the sodium and chloride content of 1 kDa low molecular weight plasma was the highest among all samples.

[0059] Table 2

[0060] sample Sodium content (%) Chlorine content (%) 1kDa low molecular weight blood cells 1.5 0.9 1kDa low molecular weight plasma 5.1 6.4 3kDa low molecular weight blood cells 1.1 0.5 3kDa low molecular weight plasma 4.2 6.7 5kDa low molecular weight blood cells 1.5 0.3 5kDa low molecular weight plasma 4.3 6.2

[0061] The sensory evaluation was designed to provide a clear and comprehensive understanding of the taste characteristics of the samples. Due to safety reasons, high molecular weight fractions were not used in the sensory experiments. The sensory samples were prepared by dissolving each low molecular weight fraction from blood cells and plasma in model broth to a final concentration of 10 g / L and adjusting the pH to 6 by adding 1% formic acid in water. The panelists were asked to evaluate six taste qualities including umami, kokumi, sweetness, saltiness, sourness and bitterness with a score ranging from 0 (not detected) to 5 (strongly detected), such as Figure 1 As shown in (a) and (b).

[0062] The experimental results showed that the 3kDa cut-off low molecular weight (LMW) of hydrolyzed chicken blood cells had significantly (P<0.01) more taste activity and taste modulation properties for kokumi (3kDa: 1.6), sweet (3kDa: 0.6) and bitter (3kDa: 2.0) tastes greater than 1kDa. Both saltiness and sweetness were unfavorable. The umami enhancement of 1kDa and 3kDa cut-off was similar. In addition, a two-three point test was performed on blood cells and plasma to compare the low molecular weight fractions of 3kDa and 5kDa cut-off. The sensory test was divided into two series, AAB and BBA (random order). The expert panelists were asked to select a sample that was different from the other two samples in each group. In order to compare the low molecular weight fractions of hydrolyzed blood cells of 3kDa and plasma of 3kDa, a two-three point test was designed to select samples with more obvious taste compared with the other two samples. According to the experimental results, there was no significant difference (P=0.1), which indicates that peptides above 3kDa did not show obvious taste activity and taste modulation activity. Compared with the low molecular weight fraction of blood cells, the 3 kDa low molecular weight plasma showed more obvious umami, kokumi and salty taste impressions. Therefore, the 3 kDa cut-off low molecular weight plasma fraction was selected for further study.

[0063] 3. Fractionation of 3 kDa low molecular weight plasma by medium pressure liquid chromatography (MPLC)

[0064] 2 g of 3 kDa LMW plasma powder was dissolved in 20 mL of deionized water and fractionated by reverse phase (RP) medium pressure liquid chromatography (Biichi, Flawil, Switzerland) on 150 x 40 mm polypropylene cassettes using 25-40 μm LiChroPrep RP18 block material (Merck, Darmstadt, Germany) as stationary phase and a gradient of 1% formic acid in water (solvent A) and methanol (solvent B) as mobile phases. The separation was monitored using an evaporative light scattering detector (ELSD) (SEDERE, Alfortville Cedex, France) and the chromatography was performed with 100% solvent A (flow rate of 40 mL / min) for 10 min, then solvent B was increased to 30% over 15 min, increased to 100% over another 15 min, and then eluted with 100% solvent B for 15 min. Finally, seven fractions (F1-F7) were collected, separated from the solvent in vacuo, and lyophilized twice before being subjected to taste dilution analysis (TDA), specific taste dilution analysis (cTDA), and chemical analysis ( Figure 2 (a), (b) and (c)).

[0065] TDA was performed to find the most intense taste-active fractions and cTDA was performed to find the taste-modulating fractions. For TDA, serial 1:2 dilutions of each fraction or subfraction were prepared in water and then presented to the sensory panelists in order of increasing concentration. Each dilution was sensory evaluated using a two- to three-point test. The dilution at which a difference in taste between the diluted fraction and two blanks (water) could be detected was defined as the taste dilution (TD) factor. The TD factors evaluated by five different panelists were averaged. The TD factors of individuals and three different sessions did not differ by more than one dilution step. A total of eight dilution steps were performed (Oliver Frank, Jezusek, & Hofmann, 2003). For cTDA, the same procedure was used as for TDA except that the fractions were dissolved in model broth and the model broth was used as the blank (Dunkel et al., 2007; Salger et al., 2019). The comparative taste dilution (cTD) factor was defined as the dilution at which a difference in taste between the diluted fraction and two blanks (model broth) could be just detected.

[0066] According to TDA and cTDA results, fraction F3 showed the highest taste activity and taste modulation properties. Figure 2 As shown in (a), in water, fraction F2 showed the highest intrinsic umami taste, with a TD coefficient of 16, followed by fractions F3 (TD coefficient = 8) and fraction F4 (TD coefficient = 4). In terms of saltiness, fractions F2 and F3 had the same TD coefficient of 4, and fraction F6 showed the highest bitter and astringent taste (TD coefficient = 4). Figure 2As shown in (b), in the model broth, fraction F3 showed the highest umami enhancing activity (cTD coefficient = 16), followed by fractions F2 (cTD coefficient = 8) and fraction F4 (cTD coefficient = 4). Fraction F4 had the highest kokumi taste. None of the fractions caused any bitterness in the model broth, which highlights the potential of fractions F2-F5 as taste enhancing fractions without unpleasant inherent taste ( Figure 2 (c)). Since fractions F2, F3, and F4 had the highest TD and cTD coefficients, further analysis focused on these fractions.

[0067] 4. Sub-fractionation of fractions F2, F3 and F4 by high performance liquid chromatography (HPEC)

[0068] To identify the taste active and taste modulating peptides in fractions F2, F3 and F4, these fractions were further separated by preparative HPLC using gradients of 1% formic acid in water and acetonitrile.

[0069] To isolate fraction F2, 10 g of fraction F2 were dissolved in 1 liter of deionized water and membrane filtered (0.45 μm). 0.5 ml of this solution was injected into an HPLC system (Jasco, Groβ-Umstadt, Germany). HPLC separation was performed under the following conditions.

[0070] Chromatographic column: Luna HILIC (200A, 250x21.2 mm, 5 μm)

[0071] Flow rate: 18 ml / min

[0072] Detector: ELSD and UV (254 nm)

[0073] To isolate fraction F3, 10 g of fraction F3 were dissolved in 1 liter of deionized water and membrane filtered (0.45 μm). 1 ml of this solution was injected into an HPLC system (Jasco, Groβ-Umstadt, Germany). HPLC separation was performed under the following conditions.

[0074] Chromatographic column: Luna PFP (2) (100A, 250x21.2 mm, 5 μm)

[0075] Flow rate: 21 ml / min

[0076] Detector: ELSD and UV (254 nm)

[0077] For the isolation of fraction F4, 20 g of fraction F4 were dissolved in 1 liter of deionized water and membrane filtered (0.45 μm). 1 ml of this solution was injected into an HPLC system (Jasco, Groβ-Umstadt, Germany). HPLC separation was performed under the following conditions.

[0078] Chromatographic column: Nucleodur C18 Pyramid (100A, 250x21.2 mm, 5 μm)

[0079] Flow rate: 20 ml / min

[0080] Detector: ELSD and UV (254 nm)

[0081] Subfractions of F2, F3 and F4 were analyzed by liquid chromatography time-of-flight mass spectrometry (LC-TOF-MS) analysis using a C8 column, and peptides were identified using PEAKS software.

[0082] Based on the experimental results, since fraction F2 contained high concentrations of known taste-active and taste-modulating molecules (such as cations, amino acids, and glutamyl dipeptides), fractions F3 and F4 were selected for large-scale separation to identify novel taste-active and taste-modulating peptides.

[0083] Example 2: Identification and evaluation of taste activity and taste modulating effects of peptides

[0084] 1. Peptide Identification

[0085] After analysis of HPLC subfractions of MPLC fractions F2, F3 and F4, potential taste-active and taste-modulating peptides in fractions F3 and F4 were identified using proteomic and sensory-omic techniques, so-called sensory proteomics.

[0086] After analyzing the HPLC subfractions of MPLC fractions F2, F3 and F4, potential taste active peptides and taste modulating peptides in fractions F3 and F4 were identified using proteomics and sensory omics techniques adapted from the methods proposed by Sebald et al., J. Agric. Food Chem. 2018, 66, 11092-11104 (i.e., the so-called sensory proteomics approach) (adapted from the methods represented by "Sebald et al., J. Argic. Food Chem. 2018, 66, 11092-11104"). Both targeted and non-targeted proteomics can be used to improve the productivity of the present invention.

[0087] By using a targeted proteomics approach, all theoretically possible peptides in chicken serum albumin and corticosteroid binding globulin were investigated and a total of 767 peptides were detected. Following non-targeted proteomics using Maxquant and PEAKS software, peptides were identified in LC-TOF-MS data of subfractions of MPLC fractions F2, F3 and F4. All peptides with an ALC (average local confidence) greater than 80 and with an N-terminal E, P or R identified by de novo sequencing and database search using PEAKS in the F3 and F4 fractions were selected. By using Maxquant, the taste active and taste modulating fractions were first investigated and the identified peptides were then classified according to the N-terminal E, P or R and peak area. Eleven peptides from fraction F3 and 44 peptides from fraction F4 were identified by MS 2 The data was further confirmed.

[0088] After the peptides were identified, several filtering steps were performed to select candidate peptides for validation. The candidate peptides finally identified in MPLC fractions F3 and F4 are shown in Table 3 below.

[0089] Table 3

[0090]

[0091]

[0092]

[0093]

[0094] Twenty-one candidate peptides were synthesized and quantitatively 1 Their purity was evaluated by H-NMR and ranged from 62% to 100%. The presence of peptides in the samples was checked by LC-MS / MS using Skyline software. The MRM (multiple reaction monitoring) transitions detected in MPLC fractions F3 or F4 were compared with the synthesized peptides to confirm that the candidate peptides were present in the hydrolyzed chicken plasma of the present invention.

[0095] 2. Determination of Taste Threshold of Peptides

[0096] The threshold concentrations of 21 newly identified peptides were determined in water (for intrinsic taste) and in model broth (adjusted to pH 6.0 with 1% formic acid) (for taste modulating properties) (Table 4). The peptides were dissolved, serially diluted in a 1:2 ratio, and then presented to the expert panel in increasing concentrations using a two- to three-point test as reported in the literature. The geometric mean of the last and penultimate concentrations was calculated and used as the individual recognition threshold. The threshold of the sensory panel was approximated as the average of the individual thresholds in two independent sessions.

[0097]

[0098] in

[0099] C p : Threshold of panel members,

[0100] C1: the first correctly evaluated concentration,

[0101] C0: The concentration of the first misjudgment

[0102]

[0103] in

[0104] C s : Thresholds for all panel members,

[0105] C p : The threshold of one panel member,

[0106] n: number of expert panel members

[0107] Table 4

[0108]

[0109]

[0110]

[0111] According to the results, the taste threshold concentration in water ranged from 100 to 1024 μmol / L. Of the 21 peptides, 7 did not show any intrinsic taste at a concentration of up to 2 mmol / L (i.e., peptides of SEQ ID NOs: 1, 6, 8, 14, 15, 17, and 19).

[0112] In addition, in the model broth, all 21 peptides showed taste modulation properties in the range of 26 μmol / L to 351 μmol / L. The peptide with SEQ ID NO:14 had the lowest umami and kokumi thresholds (26 μmol / L). The peptide with SEQ ID NO:8 had kokumi and salty enhancing properties, with the highest threshold of 351 μmol / L. The peptides with SEQ ID NO:3, 7 and 20 had kokumi, umami and salty modulating properties, with thresholds of 236, 95 and 61 μmol / L, respectively. The peptides with SEQ ID NO:12, 17, 18, 19 and 20 showed kokumi and umami enhancing properties, with thresholds of 137, 101, 247, 212 and 61 μmol / L, respectively.

[0113] In addition, peptides having SEQ ID Nos: 1, 3, 6, 8, 11, 13, 14, 16 and 21 have salty taste modulating properties and were selected for salt isointensity testing to evaluate their saltiness enhancing or reducing properties, which will be further described.

[0114] 3. Salt strength test of peptides

[0115] To investigate the effect of the peptides on the perception of salty taste, a salt isointensity test was performed according to the literature. In the salt isointensity test, the peptides (1 mmol / L) were added to a model broth containing 50 mmol NaCl, and the panelists were asked to evaluate the saltiness intensity compared to the following NaCl concentrations (30, 40, 50, 60, 70, and 80 mmol / L). The results are shown in Table 5.

[0116] Table 5

[0117]

[0118] In the model broth, the perceived salt isointensity of all peptides ranged from 42.9 to 66.7 mmol / L NaCl. Peptides with SEQ ID NOs: 6, 8, 11, 13, 14, 16 and 21 showed salty taste enhancing properties, while peptides with SEQ ID NOs: 1, 3 and 13 showed salty taste reducing properties. The strongest salt-increasing effect was observed with the peptide with SEQ ID NO: 8, with an increase of 33% (from 50 to 66.7 mmol / L of perceived salt isointensity), while the lowest salt-increasing effect was observed with the peptide with SEQ ID NO: 21, with an increase of 20% (from 50 to 59.8 mmol / L of perceived salt isointensity). In addition, salt-reducing effects were observed with the peptides with SEQ ID NOs: 1, 3 and 13, with 14% and 13%, respectively.

[0119] 4. Identification of key taste-active peptides and taste-modulating peptides

[0120] To identify peptides that play a key role in taste activity and taste modulation, not only the newly identified 21 peptides, but also the basic tastants known in the literature (such as amino acids, γ-glutamyl peptides, cations, anions, etc.) were quantified in hydrolyzed chicken plasma samples. The taste quality, taste threshold, dose over threshold (DoT) coefficient and concentration of the peptides in hydrolyzed chicken plasma samples for the 21 newly identified peptides are shown in Table 6. The taste thresholds determined for the newly identified peptides (which are listed in Table 6) were used to calculate the DoT coefficient, which is defined as the ratio of the concentration of a substance to its corresponding taste threshold concentration. The DoT coefficient can provide a preliminary estimate of the taste contribution of the taste, because substances with DoT coefficients ≥ 1 are most likely to contribute directly to the perceived taste.

[0121] Table 6

[0122]

[0123]

[0124] The results showed that five peptides with DoT coefficients greater than 1 (SEQ ID NOs: 3, 6, 11, 14 and 15) played a key role in the taste activity and taste modulation effects in plasma samples.

[0125] Example 3: Taste modulating effects of identified peptides in food

[0126] In order to investigate the flavoring properties of the identified peptides in food, further sensory experiments were performed using commercial chicken broth and model broth. According to the sensory experiment, 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 added to commercial chicken broth (Netto Marken-Discount AG & CO. KG, Maxhütte-Haidhof) and model broth. According to the experiment, 3 grams of commercial chicken broth powder were dissolved in 150 ml of boiling water. The peptides (2 mmol / l, respectively) were added to 20 mL of this solution. Afterwards, the taste threshold was determined by a two-three point test as described above (with chicken broth as reference). Table 7 shows the sensory evaluation of the identified peptides in commercial chicken products and model broth.

[0127] Table 7

[0128]

[0129] From the results, peptides with SEQ ID NO: 1, 3 and 7 affect the taste characteristics of commercial chicken broth at about 1500 μmol / L. If the peptide has SEQ ID NO: 5, the added samples cannot be distinguished (the maximum concentration tested is 2 mmol / L). In commercial chicken broth, the peptide with SEQ ID NO: 2 can be detected at a concentration of 418 μmol / L, while in the model broth, this threshold is reduced by 4 times (112 μmol / L). Based on these experimental results, it can be concluded that the peptides with specific amino acid sequences according to the present invention can enhance the taste characteristics of commercial food products, although higher concentrations are required compared to the model broth.

[0130] In addition, further commercial foods with different matrices such as chicken soup (Netto Marken-Discount AG & CO. KG, Maxhütte-Haidhof), vegetable soup (Hügli Nahrungsmittel GmbH, Radolfzell) and chili sauce (Unilever Deutschland, Hamburg) purchased from local supermarkets were used to evaluate the taste modulating effect of the novel peptides identified according to the present invention, such as 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). The food samples were prepared according to the detailed instructions listed on the different products. For chicken broth and vegetable broth, 3 grams of sample were dissolved in 150 mL of boiling water and then cooled to room temperature in an ice bath. For chili sauce, 4 grams of sample were dissolved in 150 mL of boiling water. Then, 200 micromol / L of peptides were added to 20 mL of broth or sauce. The 3-AFC test was designed to detect the taste properties of the peptides in each commercial product: two samples contained only 20 mL of the commercial product and one sample was 20 mL of the commercial product with the peptides added. The expert panel was asked to choose a different sample. Table 8 shows the results of the sensory testing of three commercial foods to which the five exemplary peptides were added.

[0131] Table 8

[0132]

[0133] The results clearly show that the novel peptides identified according to the present invention can be used as taste modifiers in edible products, in particular foods or even pharmaceuticals.

[0134] Example 4: Effect of enzyme type on peptide sequence

[0135] In order to investigate the effect of enzyme type on the amino acid sequence of peptides in hydrolyzed plasma samples, chicken plasma was digested with different enzymes such as flavorin and thermolysin.

[0136] For flavor protease digestion, 0.0504 g of chicken plasma was extracted three times with methanol and the samples were dried under nitrogen. The samples were redissolved in 1 M TEAB, urea was added to a final concentration of 8 M, and then 100 mM DTT was added and stored at 56°C for 45 minutes, followed by 0.55 M IAA and incubation at room temperature in the dark for 1 hour. The samples were then digested with flavor protease at 37°C for 15 or 30 minutes. Concentrated formic acid was added to stop the digestion, and the peptide mixture was purified by solid phase extraction on a C18 cartridge.

[0137] For thermolysin digestion, plasma samples were first prepared by adjusting the pH to 8 and then incubating 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.

[0138] Identification of peptides was performed by de novo sequencing of LC-TOF-MS data using PEAKS as previously described for screening 21 peptides (SEQ ID NO: 1-21, Table 3) in hydrolyzed chicken plasma samples. Chicken plasma was digested with flavor protease for 15 or 30 min due to the high activity of flavor protease.

[0139] Table 9 shows exemplary peptides identified in chicken plasma after flavor protease digestion, and Table 10 shows exemplary peptides identified in chicken plasma after thermolysin digestion.

[0140] Table 9

[0141]

[0142]

[0143] In the case of flavor protease digestion, it was found that the length of peptides identified by 15-minute digestion was higher than that by 30-minute digestion. The longer the digestion time, the more thorough the digestion, thus generating a greater number of short peptides.

[0144] Table 10

[0145]

[0146] Furthermore, targeted LC-MS / MS analysis of 21 peptides newly identified according to the present invention (SEQ ID NOs: 1-21, Table 3) in the digests of flavorin and thermolysin showed that none of these peptides were present due to different enzyme behaviors.

Claims

1. An isolated peptide, the amino acid sequence of which is 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).

2. The isolated peptide according to claim 1, which imparts at least one of the following effects: salt reduction, salt increase, umami and kokumi effects.

3. The isolated peptide according to claim 1 or 2, which confers a salt-reducing effect and 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).

4. The isolated peptide according to claim 1 or 2, which confers a salt-increasing effect and 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).

5. The isolated peptide according to claim 1 or 2, which imparts an umami effect and 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).

6. The isolated peptide according to claim 1 or 2, which imparts a kokumi effect and 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).

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

8. The isolated peptide according to claim 7, which is 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 performed by using a serine protease.

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

12. Use of an isolated peptide as defined in any one of claims 1 to 11 as a taste modifier for food or pharmaceutical preparations.

13. 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 A combination of them.

14. The protein hydrolysate according to claim 13, comprising at least one peptide having the following amino acid sequence: 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).

15. The protein hydrolysate according to any one of claims 13 or 14, which is obtained by enzymatic hydrolysis.

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

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

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

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

20. Use of a protein hydrolysate as defined in any one of claims 13 to 19 as a taste modifier for food or pharmaceutical preparations.

21. A taste modifying composition comprising an isolated peptide as defined in any one of claims 1 to 11.

22. A taste modifying composition comprising a protein hydrolysate as defined in any one of claims 13 to 19.

23. Use of the taste-adjusting composition according to claim 21 or 22 in food or pharmaceutical preparations.

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

25. A food preparation comprising a protein hydrolysate as defined in any one of claims 13 to 19.

26. A food preparation comprising a taste modifying composition as defined in claim 21 or 22.

27. A pharmaceutical preparation comprising an isolated peptide as defined in any one of claims 1 to 11.

28. A pharmaceutical preparation comprising a protein hydrolysate as defined in any one of claims 13 to 19.

29. A pharmaceutical formulation comprising a taste modifying composition as defined in claim 21 or 22.

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

31. A method for adjusting the taste of a food preparation, comprising applying an effective amount of a protein hydrolysate as defined in any one of claims 13 to 19 and comprising an effective amount of isolated peptides to the food preparation.

32. A method for adjusting the taste of a food preparation, comprising applying a taste-modulating composition as defined in claim 21 or 22 comprising an effective amount of an isolated peptide to the food preparation.

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

34. A method for adjusting the taste of a pharmaceutical preparation, comprising applying a protein hydrolyzate as defined in any one of claims 13 to 19 and comprising an effective amount of isolated peptides to the pharmaceutical preparation.

35. A method for adjusting the taste of a pharmaceutical preparation, comprising applying a taste-modulating composition as defined in claim 21 or 22 comprising an effective amount of an isolated peptide to the pharmaceutical preparation.

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