Taste-improving peptide

Abstract

To provide a milk-derived peptide having a taste-improving effect, a taste-improving agent comprising the same, and a food or drink containing the same. A taste-improving agent comprising a peptide, said peptide having the sequence(s) Val-Pro and/or Leu-Leu-Leu, and a food or drink to which the taste-improving agent is added.

Claims

1. A method for improving the aftertaste of an artificial sweetener, comprising: Adding a taste-improving agent comprising a peptide having the sequence Val-Pro and a peptide having the sequence Leu-Leu-Leu to a drink comprising the artificial sweetener, wherein the mass ratio of Val-Pro:Leu-Leu-Leu is 9:1 to 3:7 in a mixture of Val-Pro and Leu-Leu-Leu, wherein the addition amount of the mixture of Val-Pro and Leu-Leu-Leu is 10 ppb to 100 ppm based on the drink comprising the artificial sweetener.

2. The method according to claim 1, wherein the drink is tea.

3. A method for enhancing the richness of milk, comprising: adding a taste-improving agent comprising a peptide having the sequence Val-Pro and a peptide having the sequence Leu-Leu-Leu to a drink comprising milk or a dairy product, wherein the mass ratio of Val-Pro:Leu-Leu-Leu is 9:1 to 3:7 in a mixture of Val-Pro and Leu-Leu-Leu, wherein the addition amount of the mixture of Val-Pro and Leu-Leu-Leu is 100 ppb to 100 ppm based on the drink comprising milk or the dairy product.

4. A taste-improving agent consisting of a peptide having the sequence Val-Pro and a peptide having the sequence Leu-Leu-Leu, wherein the mass ratio of Val-Pro:Leu-Leu-Leu is 9:1 to 3:7 in a mixture of Val-Pro and Leu-Leu-Leu.

5. A food or a drink to which the taste-improving agent according to claim 4 is added, wherein the addition amount of the mixture of Val-Pro and Leu-Leu-Leu is 10 ppb to 100 ppm based on the food or the drink.

Description

EXAMPLES

(1) Hereinafter, the present invention will be described in detail by way of Examples.

[Example 1] (Preparation and Identification of Peptide)

(2) 1. Production of Peptide Mixture by Enzymatic Decomposition

(3) To 70 g of ion exchanged water, 24 g of TMP (H) (Total Milk Protein, produced by MORINAGA MILK INDUSTRY CO., LTD.) was dissolved and sterilized by heating. Then, the enzyme solution obtained by dissolving 0.5 g of protease A2G, 0.1 g of protease NG and 0.3 g of Flavourzyme 1000 L in 6 mL of water was added to the solution, and the enzyme reaction was carried out at 32 C. for 24 h while stirring. After that, the reaction mixture was heat-inactivated to obtain a peptide mixture.

(4) 2. Fractionation of Peptide Mixture by UF Membrane

(5) The peptide mixture obtained in the above procedure 1 was centrifuged (20,000 G) to obtain an aqueous layer. The aqueous layer was treated with UF membranes of 20 nm, 5 kDa and 1 kDa, respectively, and permeate solutions were collected. Each permeate solution obtained through the UF membranes or the peptide mixture before treatment were added to a commercially available yoghurt drink. After the yoghurt drink was stored at 10 C. for 10 days, five special panelists conducted sensory evaluation regarding a sourness-masking effect. Note that each sample was prepared such that the addition amount is 0.1% calculated in terms of the peptide mixture. A commercially available yoghurt drink without addition was used as control. Evaluation was performed by scoring from 0 to 3 shown in Table 1 at 0.5 point intervals.

(6) TABLE-US-00001 TABLE 1 Evaluation Score Evaluation 0 No effect 1 A little effect 2 Clear effect 3 Highest effect

(7) This result is shown in the following Table 2.

(8) TABLE-US-00002 TABLE 2 Sample Name Evaluation Result No Addition 0.0 Peptide Mixture 2.1 20 nm Membrane Permeate Solution 2.2 5 kDa Membrane Permeate Solution 2.0 1 kDa Membrane Permeate Solution 2.3

(9) From this result, it was confirmed that the 1 kDa membrane permeate solution has the same or stronger taste-improving effect than the peptide mixture before treatment.

(10) 3. Fractionation of 1 kDa Membrane Permeate Solution by HPLC

(11) From the above-mentioned results of sensory evaluation, the 1 kDa membrane permeate solution was further fractionated by HPLC. The 1 kDa membrane permeate solution was fractionated into 13 fractions from A fraction to M fraction under the following conditions.

(12) Column: ODS-2 (0 20250 mm)

(13) Eluent: water/ethanol

(14) Flow rate: 3.0 mL/min.

(15) Detector: UV (220 nm)

(16) Temp: 40 C.

(17) In the same manner as in the above procedure 2, the obtained 13 fractions were added to a commercially available yoghurt drink, respectively. After the yoghurt drink was stored at 10 C. for 10 days, five special panelists conducted sensory evaluation regarding a sourness-masking effect. Each sample was prepared such that the addition amount is 0.1% calculated in terms of the peptide mixture. The evaluation method is the same as in the above procedure 2. As a result, it was confirmed that the fraction E has the same high taste-improving effect as the peptide mixture obtained in the above procedure 1.

(18) 4. This Fraction was Further Subjected to HPLC (Gel Filtration Column) Fractionation Under the Following Conditions to Divide into Four Fractions from E-1 Fraction to E-4 Fraction.

(19) Column: Shodex SB401-4E (4.6 mm I.D.250 mL)

(20) Eluent: H.sub.2O

(21) Flow rate: 0.2 mL/min.

(22) Detector: UV (220 nm)

(23) Temp: 30 C.

(24) In the same manner as in the above procedure 2, the obtained four fractions were added to a commercially available yoghurt drink, respectively. After the yoghurt drink was stored at 10 C. for 10 days, five special panelists conducted sensory evaluation regarding a sourness-masking effect. Each sample was prepared such that the addition amount is 0.1% calculated in terms of the peptide mixture. The evaluation method is the same as in the above procedure 2. As a result, it was confirmed that the fraction E-3 has the same high taste-improving effect as the peptide mixture obtained in the above procedure 1.

(25) 5. Structural Analysis of Peptide

(26) The fraction obtained in the above procedure 4 was lyophilized and the peptides included therein were identified. Based on the results of LCMS-IT-TOF RP-ESI-MS/MS under the conditions shown below, Manual de novo sequencing was conducted.

(27) As a result, the following five kinds of peptide were identified as major components:

(28) Val-Pro-Pro (Hereinafter, it is described as VPP in single letter code);

(29) Leu-Pro-Pro (Hereinafter, it is described as LPP in single letter code);

(30) Val-Pro (Hereinafter, it is described as VP in single letter code);

(31) Leu-Leu-Leu (Hereinafter, it is described as LLL in single letter code);

(32) Leu-Leu-Leu-Leu (Hereinafter, it is described as LLLL in single letter code); and

(33) phenylalanine (It is described as F in single letter code).

(34) Column: ODS3 (2 mm i.d., 150 mm, 3 um)

(35) Gradient: A: 0.1% Formic acid, B: Acetonitrile

(36) Auto MS/MS range: 200 to 800 Da

(37) CID energy: 80 to 100%

[Example 2] (Evaluation of Taste-Improving Effect of Peptide)

(38) The five kinds of peptide identified in Example 1 and phenylalanine were added to a commercially available yoghurt drink, respectively. After the yoghurt drink was stored at 10 C. for 10 days, five special panelists conducted sensory evaluation regarding a sourness-masking effect. The peptide mixture obtained in Example 1-1 was added in an amount of 0.1%, the peptides and phenylalanine obtained in Example 1-5 were added in an amount of 20 ppb and the mixture of VP+LLL was added in an amount of 10 ppb each. The evaluation method was the same as in Example 1-2.

(39) The results are shown in the following Table 3.

(40) TABLE-US-00003 TABLE 3 Sample Name Evaluation Result Peptide Mixture 2.4 LPP 0.3 VP 1.3 LLLL 0.3 LLL 1.4 VPP 0.2 F 0.1 VP + LLL 2.3

(41) From this result, the two kinds of peptide: VP and LLL, were confirmed to have a remarkable sourness-masking effect. Further, the same high taste-improving effect as the peptide mixture was confirmed by combining these two kinds of peptide.

[Example 3] (Minimum Addition Amount of Peptide)

(42) VP and LLL were mixed in the mixture ratio within the range from 1:9 to 9:1 and each mixture was added to a commercially available yoghurt drink in an amount of 0.01 ppm. After the yoghurt drink was stored at 10 C. for 10 days, five special panelists conducted sensory evaluation regarding a sourness-masking effect. The evaluation method is the same as in the Example 1-2. The results are shown in the following Table 4.

(43) TABLE-US-00004 TABLE 4 Sample Name Evaluation Result Peptide Mixture 2.4 VP 1.4 VP:LLL = 9:1 2.1 VP:LLL = 7:3 2.4 VP:LLL = 5:5 2.4 VP:LLL = 3:7 2.1 VP:LLL = 1:9 1.6 LLL 1.3

(44) From this result, it was confirmed that VP and LLL each may be used solely, however, inclusion of VP and LLL with the mixture ratio of 9:1 to 3:7 especially generated a synergistic effect and exhibited a higher sourness-masking effect than each peptide alone.

[Example 4] (Minimum Addition Amount of Peptide)

(45) VP, LLL and the mixture of equal parts of VP and LLL were added to a commercially available yoghurt drink in an amount of 0.001 ppm, 0.01 ppm and 0.1 ppm, respectively. After the yoghurt drink was stored at 10 C. for 10 days, five special panelists conducted sensory evaluation regarding a sourness-masking effect. The evaluation method is the same as in the Example 1-2. The results are shown in the following Table 5.

(46) TABLE-US-00005 TABLE 5 Addition Concentration Evaluation Sample Name [ppm] Result Peptide Mixture 1000 2.4 LLL 0.001 0.3 LLL 0.01 1.4 LLL 0.1 1.5 VP 0.001 0.2 VP 0.01 1.5 VP 0.1 1.5 VP:LLL = 1:1 0.001 0.5 VP:LLL = 1:1 0.01 2.3 VP:LLL = 1:1 0.1 2.4

(47) From this result, it was confirmed that all of VP alone, LLL alone and the mixture of equal parts of VP:LLL have a high sourness-masking effect with the concentration of 0.01 ppm or more.

[Example 5] (Aftertaste-Improving Effect of Artificial Sweetener)

(48) To a commercially available black tea beverage (straight tea) using artificial sweeteners (xylitol, acesulfame K and sucralose), the peptide mixture and the mixture of equal parts of VP and LLL were added. Five special panelists conducted sensory evaluation regarding an aftertaste-improving effect of a sweet taste. The peptide mixture was added in an amount of 0.02% and the mixture of equal parts of VP and LLL was added in an amount of 0.001 ppm, 0.01 ppm and 0.1 ppm, respectively. The evaluation method was the same as in Example 1-2. The results are shown in the following Table 6.

(49) TABLE-US-00006 TABLE 6 Concentration Addition Component [ppm] Evaluation Result No Addition 0.0 Peptide Mixture 200 2.3 VP:LLL = 1:1 0.001 0.9 VP:LLL = 1:1 0.01 2.2 VP:LLL = 1:1 0.1 2.1

(50) As a result, it was confirmed that the mixture of equal parts of VP and LLL has a remarkable aftertaste-improving effect of the sweet taste of an artificial sweetener with the concentration of 0.01 ppm or more.

[Example 6] (Enhancing Effect of Richness of Milk)

(51) The peptide mixture and the mixture of equal parts of VP and LLL were added to a commercially available tea with milk, and five special panelists conducted sensory evaluation regarding an enhancing effect of the richness of milk. The peptide mixture was added in an amount of 0.10% and the mixture of equal parts of VP and LLL was added in an amount of 0.01 ppm, 0.1 ppm and 1 ppm, respectively. The evaluation method was the same as in the Example 1-2. The results are shown in the following Table 7.

(52) TABLE-US-00007 TABLE 7 Concentration Addition Component [ppm] Evaluation Result No Addition 0.0 Peptide Mixture 1000 2.5 VP:LLL = 1:1 0.01 1.0 VP:LLL = 1:1 0.1 2.4 VP:LLL = 1:1 1.0 2.6

(53) As a result, it was confirmed that the mixture of equal parts of VP and LLL has a remarkable enhancing effect of the richness of milk with the concentration of 0.1 ppm or more.

INDUSTRIAL APPLICABILITY

(54) The present invention provides a taste-improving agent comprising a specific di- or tripeptide, which is useful for improving the taste of various foods, especially health foods and foods containing an artificial sweetener, and therefore, has applicability in the food industry.