Umami enhancer for food

Abstract

To provide novel technology capable of enhancing umami in food materials.SOLUTION: An umami enhancer for food comprises an extract of Lentinus edodes or Flammulina velutipes and is brought into contact with food in the presence of a chloride salt.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to an umami enhancer for food. [Background technology]

[0002] Monosodium glutamate is an amino acid known to exhibit a flavor termed umami, and is used to enhance the umami taste of foods. It is also mass-produced artificially by microbial fermentation around the world. However, it has been pointed out that excessive intake of monosodium glutamate can have side effects such as inducing diabetes and liver toxicity (Non-Patent Document 1). Therefore, ingesting a large amount of monosodium glutamate is not preferable from the viewpoint of safety. [Prior art documents] [Non-patent literature]

[0003] [Non-Patent Document 1] Zehra Kazmi, Iffat Fatima, Shaghufta Perveen & Saima Shakil Malik (2017) Monosodium glutamate: Review on clinical reports, International Journal of Food Properties, 20:sup2, 1807-1815, Summary of the Invention [Problem to be solved by the invention]

[0004] An object of the present invention is to provide a new technique capable of enhancing the umami flavor in food materials. [Means for solving the problem]

[0005] The present inventors have found that the umami taste of food is enhanced by contacting an extract of shiitake mushroom or enoki mushroom in the presence of a chloride salt with the food, and have completed the present invention.

[0006] The present invention relates to the following (1) to (4). (1) A food umami enhancer comprising an extract of shiitake mushroom or enoki mushroom, which is contacted with food in the presence of a chloride salt. (2) The umami enhancer according to (1), wherein the contact is carried out in the presence of 0.1% or more and 20% or less of the chloride salt. (3) The umami enhancer according to (1), wherein the chloride salt is selected from the group consisting of sodium chloride, potassium chloride, magnesium chloride, ammonium chloride, and calcium chloride. (4) A method for enhancing the umami taste of a food product comprising contacting an extract of shiitake or enokitake mushroom with the food product in the presence of a chloride salt. Effect of the Invention

[0007] According to the present invention, it is possible to provide a food umami enhancer containing an extract of shiitake mushroom or enoki mushroom. [Brief description of the drawings]

[0008] [Figure 1] Figure 1 shows the glutamic acid gain or content in whole shiitake mushroom extracts. Figure 1A shows the effect of NaCl on glutamic acid gain upon incubation at 40°C for 120 minutes. Figure 1B shows the effect of temperature on glutamic acid content upon incubation of shiitake mushroom extracts for 120 minutes. [Diagram 2]Figures 2A-D show the effect of NaCl on various enzymes in extracts of shiitake mushrooms (pilus, stipe or whole mushrooms). Figure 2A shows protease activity at pH 6. Figure 2B shows Glu-pNA hydrolysis (GAPase) activity at pH 6. Figure 2C shows γ-Glu-pNA hydrolysis activity (γGAPase) of the pillus and stalk and glutamic acid production activity of the whole mushroom. The pillus and stalk were measured for γ-glutamyltranspeptidase (transferase) activity, and the whole mushroom was measured for glutamic acid production activity from glutathione. Figure 2D shows glutamic acid production from 1% glutamine (glutaminase activity). [Diagram 3] Figure 3A shows the effect of NaCl on glutaminase activity in extracts of Lentinula edodes (cap or stem). Figure 3B shows the effect of NaCl on glutaminase activity in extracts of various mushrooms. Activity measurements were performed at pH 7. [Figure 4] Figure 4 shows the profile of glutaminase activity in whole shiitake mushroom extracts. Figure 4A shows the effect of pH. Figure 4B shows the effect of temperature. The activities in Figures 4A-4B are relative activities, with the highest activity in the presence of 2% NaCl defined as 100%. Figure 4C shows the effect of 0.17M chloride or sodium salt. Figure 4D shows the effect of NaCl on glutaminase activity in dried shiitake mushroom extracts. [Diagram 5] Figure 5 shows the glutamic acid content of sample extracts obtained by extracting various kinds of dried shiitake mushrooms (whole) in the presence of 20 volumes of distilled water or 1% NaCl for 23 hours at 4°C, filtering the extracts, incubating the dried shiitake mushroom extracts at 50°C for 60 minutes, stopping the reaction in a boiling water bath for 5 minutes, and centrifuging (16162 x g for 5 minutes). [Figure 6] Figure 6 shows the glutamic acid content in various foods using whole shiitake mushroom extracts supplemented with 1% NaCl. The glutamic acid content was measured after incubation at 50°C for 60 minutes. "Alone" shows the results of the extract of each food listed on the horizontal axis alone, and "Mixed" shows the results of the mixture of each food listed on the horizontal axis and shiitake mushroom extract. [Figure 7] 7A and B show the increase in glutamate after adding 0, 1, 5, or 10% NaCl to a mixture of shiitake mushroom and garlic or onion extract and incubating at 50° C. for 60 minutes. [Figure 8] Figure 8 shows the change in glutamic acid and glutamine concentrations over time in a mixture of shiitake and garlic extracts in the presence or absence of NaCl, Figure 8A was performed in the absence of NaCl, and Figure 8B was performed in the presence of 2% NaCl. [Figure 9] FIG. 9 shows the results of Butyl-650M hydrophobic chromatography of an extract of Enokitake mushroom. [Figure 10] FIG. 10 shows the GAP specific activity etc. in each purified fraction of the Enokitake mushroom extract. [Figure 11] FIG. 11 shows the results of Native-PAGE of the 2nd DEAE fraction of the Enokitake mushroom extract. [Figure 12] FIG. 12 shows the pH profile of GAP activity of partially purified enzyme from Enokitake mushroom extract. [Figure 13] FIG. 13 shows the temperature characteristics of GAP activity of a partially purified enzyme from an extract of Enokitake mushroom. [Figure 14] FIG. 14 shows the substrate specificity of the partially purified enzyme from Enokitake mushroom extract and the PAP activity at each stage of enzyme purification. [Figure 15] FIG. 15 shows the results of Native-PAGE and SDS-PAGE of the gel filtration HPLC separated peaks of the Enokitake mushroom extract. [Figure 16] FIG. 16 shows the effect of NaCl on the GAP and PAP activities of partially purified enzymes from Enokitake mushroom extracts. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0009] DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS The following embodiments of the present invention are merely examples for explaining the present invention, and are not intended to limit the present invention to the embodiments.

[0010] The present invention relates to an umami enhancer for foods, comprising an extract of shiitake mushroom or enoki mushroom.

[0011] 1. Shiitake or Enokitake mushroom extract "Shiitake" refers to Lentinula edodes, which belongs to the genus Lentinula in the family Omphalotaceae in the order Agaricales in the class Agaricomycetes in the phylum Eumycota, and "Enokitake" refers to Flammulina velutipes, which belongs to the genus Flammulina in the family Physalacriaceae in the order Agaricales in the same class. Shiitake and enokitake may be available as edible shiitake and edible enokitake.

[0012] The extract of shiitake or enokitake may be in the form of a dried and ground powder or in the form of a liquid.

[0013] The extract of shiitake or enokitake may be an extract of the pileus, stipe, or whole of shiitake or enokitake, but for shiitake extract, it is preferable to use an extract of the stipe.

[0014] The extract of shiitake or enokitake may be prepared by a person skilled in the art using a known method as appropriate, and may be, for example, a supernatant obtained by stirring shiitake or enokitake with a near-neutral buffer solution (which may be a phosphate buffer solution in some cases; the same applies below) or distilled water and subjecting the mixture to centrifugation, or an extract obtained by extraction using an organic solvent, which may be further concentrated or purified by column chromatography or molecular sieves. When preparing the extract of shiitake or enokitake, raw shiitake or enokitake may be used, or dried shiitake or enokitake may be used. The extract of shiitake or enokitake is not particularly limited, and may be a supernatant obtained by mixing 0.5 to 20 times, 0.5 to 5 times, 0.5 to 4 times, 0.8 to 3.2 times, about 1 to about 3 times, preferably about 3 times the mass of shiitake or enokitake with a near neutral buffer, saline or distilled water, homogenizing the mixture, and centrifuging the mixture. The supernatant may be freeze-dried into a powder form to be used as the extract of shiitake or enokitake. Homogenization may be performed by an alternative means capable of pulverizing shiitake or enokitake.

[0015] The amount of the extract of shiitake or enokitake mushrooms when contacting with food is not particularly limited as long as it has the effect of enhancing the umami taste of the food, but may be, for example, 0.1% by mass or more, 0.25% by mass or more, 0.5% by mass or more, 0.75% by mass or more, 0.1% by mass or more, 20% by mass or less, 18% by mass or less, 16% by mass or less, 14% by mass or less, 12% by mass or less, or 10% by mass or less, relative to 100% by volume of the food, calculated as the above-mentioned supernatant liquid. In other words, the amount of the extract of shiitake or enokitake mushrooms may be 0.1 times or more and 2 times or less, 0.5 times or more and 1.5 times or less, or about the same amount, relative to the amount of the food, calculated as the above-mentioned supernatant liquid.

[0016] The temperature of the environment where the shiitake or enokitake extract comes into contact with the food is not particularly limited as long as it has the effect of enhancing the umami taste of the food, and may be, for example, 30° C. to 60° C., 40° C. to 60° C., or 35° C. to 40° C. to 55° C., or 50° C. or less. Although not particularly limited, when a shiitake extract is used, the temperature may be 35° C. to 50° C., or 35° C. to 45° C., and when a enokitake extract is used, the temperature may be 35° C. to 55° C., 40° C. to 55° C., or 45° C. to 55° C.

[0017] The pH of the environment in which the extract of Shiitake or Enokitake mushroom comes into contact with food is not particularly limited as long as it has the effect of enhancing the umami taste of the food, and may be 5 or more, 5.5 or more, 5.6 or more, 5.7 or more, 5.8 or more, 5.9 or more, 6.0 or more, 8.0 or less, 7.5 or less, 7.0 or less, 6.9 or less, 6.8 or less, 6.7 or less, 6.6 or less, or 6.5 or less.

[0018] The time for which the extract of shiitake or enokitake is brought into contact with the food is not particularly limited.

[0019] 2. Chloride salts In this embodiment, the "chloride salt" is not particularly limited as long as it is a metal salt that generates chloride ions when contacted with an extract of shiitake mushrooms or enoki mushrooms, but it is a salt of chloride ions and a metal. Examples of chloride salts include sodium chloride, potassium chloride, magnesium chloride, ammonium chloride, and calcium chloride. For example, in this embodiment, when sodium chloride is used as a chloride salt, this includes using sodium chloride in the form of sodium chloride salt, and using sodium ions and chloride ions dissolved by moisture contained in food or moisture present during the cooking process.

[0020] In this embodiment, "contacting the shiitake or enokitake extract with the food in the presence of a chloride salt" means that the chloride salt or its ionic form is present in the environment in which the shiitake or enokitake extract comes into contact with the food.

[0021] The amount of chloride salt present in the environment where the extract of shiitake or enokitake mushroom comes into contact with food, relative to the food containing the extract of shiitake or enokitake mushroom and chloride salt, may be 0.01 w / v% or more and 30 w / v% or less, 0.1 w / v% or more and 20 w / v% or less, 0.5 w / v% or more and 10 w / v% or less, or 1 w / v% or more and 5 w / v% or less, for example, 0.05 w / v% or more, 0.1 w / v% or more, 0.1 w / v% or more, 0.5 w / v% or more, 0.8 w / v% or more, 0.9 w / v% or more, or 1 w / v% or more, or may be 25 w / v% or less, 20 w / v% or less, 15 w / v% or less, 10 w / v% or less, 8 w / v% or less, 5 w / v% or less, 4 w / v% or less, or 3 w / v% or less. "w / v%" refers to the mass % of chloride salt relative to the total volume of the food product including shiitake or enokitake extract and chloride salt. In this embodiment, the umami taste of food is enhanced by the extract of shiitake mushroom or enokitake mushroom in the presence of chloride salt in the food. The chloride salt in the food may be the chloride salt originally present in the food, or may be the chloride salt present in the food by adding chloride salt to the food. In the food in which chloride salt is present, the umami taste is enhanced by contacting the extract of shiitake mushroom or enokitake mushroom with the food. Therefore, when the food umami enhancer of this embodiment is used, both the chloride salt and the extract of shiitake mushroom or enokitake mushroom are present in the food, that is, they coexist. The presence of chloride salt can be expressed as the content ratio of the chloride salt in mass % relative to the entire food, and it is preferable that the chloride salt is present at 0.01 w / v% or more and 30 w / v% or less. The amount of chloride salt present may be set within the above numerical range described in this paragraph. In this embodiment, the food may contain a predetermined mass % of shiitake or enokitake extract relative to 100 mass % of the food in the presence of a predetermined amount of chloride salt.

[0022] 3. Umami Umami is a type of flavor, and is one of the flavors exhibited by substances such as glutamic acid and ribonucleotides such as inosinic acid (5'-IMP), adenosinic acid (5'-AMP), and guanosinic acid (5'-GMP). Umami substances that exhibit umami can be produced by decomposition reactions of proteins, amino acids, or nucleic acids during food processing, including aging and fermentation. Thus, the umami of a food can be enhanced by increasing the amount of the umami substance, increasing the activity of an enzyme that produces the umami substance, and the like. In this embodiment, "enhancing the umami of a food" may mean an increase in the amount of the umami substance in the food, or an increase in the activity of an enzyme that produces the umami substance in the food.

[0023] The umami of a food can be measured by the activity of an enzyme that decomposes proteins, amino acids, or nucleic acids to produce an umami substance, or the amount of the umami substance. The amount of an umami substance may be measured by a method known to those skilled in the art, as appropriate, for example, in the case of glutamic acid, the amount may be measured using an L-glutamic acid measurement kit, Yamasa (NEO), or the like. The activity of an enzyme that produces an umami substance may be measured by a method known to those skilled in the art, as appropriate, for example, the activity of a protease, peptidase, glutaminase, or the like, which is known as such an enzyme, may be measured.

[0024] The food umami enhancer of this embodiment includes an extract of shiitake mushroom or enokitake mushroom, and enhances the umami of the food by contacting the food with the food in the presence of a chloride salt. That is, the food umami enhancer of this embodiment includes an extract of shiitake mushroom or enokitake mushroom. When the food umami enhancer is added to the food, it is preferable that a chloride salt is present in the food. The chloride salt may be added after the food umami enhancer of this embodiment is added, or the chloride salt may be added to the food first and then the food umami enhancer of this embodiment is added, or the chloride salt present in the food may be used to add the food umami enhancer of this embodiment to the food. Therefore, the food umami enhancer of this embodiment contains an extract of Shiitake or Enokitake mushroom, and the umami of the food is enhanced by the coexistence of the chloride salt and the extract of Shiitake or Enokitake mushroom in the food. This enhancing effect is thought to be due to the action of enzymes that produce umami substances contained in the extract of Shiitake or Enokitake mushroom in the presence of the chloride salt, but the cause is not limited to this. Contacting with food in the presence of a chloride salt means that the extract of Shiitake or Enokitake coexists with the food in the presence of a chloride salt, and may mean a mixture of the extract of Shiitake or Enokitake, the chloride salt, and the food. In the case where the chloride salt is originally contained in the food, it may be a mixture of the extract of Shiitake or Enokitake and the food that originally contains the chloride salt. The chloride salt may be a chloride salt derived from the food, or may be a chloride salt added to the food. The chloride salt may be added as a chloride salt contained in a seasoning. In this embodiment, the composition may be a composition containing an extract of shiitake mushrooms or enokitake mushrooms, in which the umami taste of the food is enhanced by the presence of a chloride salt and an extract of shiitake mushrooms or enokitake mushrooms in the food. Therefore, in this embodiment, the composition may be a composition containing an extract of shiitake mushrooms or enokitake mushrooms, in which the umami taste of the food is enhanced by the coexistence of the extract of shiitake mushrooms or enokitake mushrooms and a chloride salt in the food.

[0025] The food is not particularly limited as long as it is a food that is capable of producing umami substances, and may be a processed or cooked food, an unprocessed or uncooked food, or a food that is in the process of being processed or cooked.

[0026] The umami enhancer of the present embodiment can be used as a food product for the purpose of enhancing the umami of the food product, and can be used in the form of, for example, granules, powder, liquid, paste, or the like. These may be used during the cooking process, before or after cooking, or when eating or drinking.

[0027] The umami enhancer of the present embodiment may contain other additive materials within the range of the effect of enhancing the umami of food, such as antioxidants such as ascorbic acid fatty acid esters, lignans, coenzyme Q, γ-oryzanol, tocopherol, flavorings, spice extracts, animal extracts, flavoring materials such as fatty acids, emulsifiers, silicones, and colorings.

[0028] The present invention also relates to a composition for enhancing the umami taste of a food by contacting the food in the presence of a chloride salt, the composition comprising an extract of shiitake or enokitake.The present invention also relates to a method for enhancing the umami taste of a food, the method comprising contacting a food in the presence of a chloride salt with an extract of shiitake or enokitake.The present invention also relates to a method for producing a food with enhanced umami taste, the method comprising contacting a food in the presence of a chloride salt with an extract of shiitake or enokitake. In the present invention, the presence of a chloride salt may be achieved by cooking with salt-koji etc. In the present embodiment, the method may be such that the umami of a food is enhanced by the presence of a chloride salt and an extract of shiitake mushroom or enoki mushroom in the food. EXAMPLES

[0029] Using the methods described below, we measured the enzyme activity and glutamic acid content in extracts from mushrooms such as shiitake and enokitake when they were brought into contact with food in the presence of chloride salts. The results are shown in Figures 1 to 16.

[0030] (Sample preparation) Fresh mushrooms such as shiitake and enokitake were purchased from a grocery store in Tokyo. Each mushroom was cut into small pieces, and three or an equal volume of distilled water was added and homogenized in a blender for 40 seconds. The mixture was then centrifuged (16,000×g, 10 minutes), and the resulting supernatant was used as the sample extract. Sample extracts were prepared from the whole shiitake mushroom, the cap, and the stalk. Dried shiitake mushrooms were prepared by grinding dried shiitake mushrooms from Miyashiro Co., Ltd. (Hyogo Prefecture) in a mill, adding 10 times the amount of distilled water, and storing the mixture in a refrigerator at 4°C for 24 hours. The mixture was then centrifuged (6570 × g, 10 minutes). The resulting supernatant was used as the dried shiitake mushroom extract. After grinding the dried shiitake mushrooms in a mill, 20 volumes of distilled water or NaCl solution were added, and the mixture was stored in a refrigerator at 4°C for 23 hours, after which it was filtered through filter paper. The filtrate was incubated at 50°C for 60 minutes, the reaction was stopped in a boiling water bath for 5 minutes, and the supernatant after centrifugation (16162 x g, 5 minutes) was used as a sample extract for free glutamic acid content. The dried shiitake mushrooms used were Koushin from Oita Prefecture (Koushin) by Kensho Co., Ltd., Kotsubu Donko from Miyazaki Prefecture (Kotsubu Donko) from Okada Shoten Co., Ltd., Kyushu Donko from Kyushu Donko from Okada Shoten Co., Ltd., and log-dried shiitake mushrooms from Shimane Prefecture (Shimane Prefecture).

[0031] (Incubation of sample extract) 1 mL of reaction solution containing 0.45 mL of sample extract was incubated in a hot water bath. The reaction was stopped by boiling water for 5 minutes, and the supernatant was centrifuged and used to measure the free glutamic acid content. To investigate the effect of NaCl, 1 mL of reaction solution was added with 2- or 5-fold concentrated NaCl solution to adjust the final concentration to the desired chloride salt concentration, and the same treatment as above was used. To investigate the free glutamic acid content in combination of sample extract and food, a mixture of equal volumes of food extract prepared in the same manner as the sample extract was mixed, to which NaCl solution was added to adjust the final chloride salt concentration to the desired w / v%.

[0032] (Measurement of glutaminase activity) Glutaminase activity was measured according to the method of Teramoto et al. (1985). 0.5 mL of distilled water or saline and 0.25 mL of sample extract were added to 1 mL of reaction solution containing 0.25 mL of 0.2 M glutamine solution (pH 7.0) as substrate solution, and the mixture was incubated at 50°C for 60 minutes. The reaction was stopped after boiling. The free glutamic acid content in the supernatant obtained by centrifugation was measured using the L-glutamic acid measurement kit Yamasa (NEO). For the blank, the same procedure was performed using 0.2 M phosphate buffer (pH 7.0) instead of the substrate solution. Glutaminase activity (U) was defined as the amount of enzyme that produces 1 μmol of glutamic acid from glutamine per minute. Britton-Robinson buffer was used as the blank substrate solution for measuring glutaminase activity at pH 4 to 8.

[0033] (Measurement of aminopeptidase activity) Glutamyl aminopeptidase activity was measured as Glu-pNA hydrolysis (GAPase) activity using Glu-pNA (Peptide Institute) as a substrate solution. Samples were measured for the increase in OD405 nm in kinetic mode on a microplate reader FlexStation 3 (Molecular Devised) together with the substrate solution. GAPase activity (U) was defined as the amount of enzyme that releases 1 μmol of nitroaniline from Glu-pNA per minute. Gamma glutamyl transpeptidase (γGAPase) activity was measured according to the method of Iwami et al. (1975), which is the same as the above method using gamma glutamyl pNA as a substrate. γGAPase activity (U) was also measured by the production of glutamic acid generated from 0.4 mM glutathione (γ-glutamyl peptide). This activity was defined as the amount of enzyme that releases 1 μmol of glutamic acid from glutathione per minute. The method used was the same as above, using glutamyl pNA as a substrate, but with glutathione as the substrate, and the amount of glutamic acid was measured using the Yamasa (NEO) L-glutamic acid measurement kit. The aminopeptidase activity in the enokitake mushroom extract was measured by measuring the increase in OD405nm. In the following, the aminopeptidase activity measured in the enokitake mushroom extract using the Glu-pNA substrate is referred to as GAP, and the aminopeptidase activity measured using the Pro-pNA substrate is referred to as PAP.

[0034] (Measurement of protease activity) The sample extract was incubated with 1.5% casein solution (pH 3.0 or 6.0) at 30°C for 10 minutes. After the reaction was stopped, it was neutralized with sodium carbonate solution, colored with phenol reagent, and OD660nm was measured. Protease activity (U) was defined as the amount of enzyme that released 1μg of tyrosine from casein per minute.

[0035] (Measurement of L-glutamic acid and L-glutamine content) The free L-glutamic acid content in the reaction solution of the sample extract was measured using an L-glutamic acid measurement kit (Yamasa NEO). The L-glutamine content in the sample extract was measured using an L-glutamine measurement kit (Enzyme-Sensor Co., Ltd.). If necessary, the concentration was calculated from the content.

[0036] (statistical analysis) Statistical analysis was performed using Excel Statistics version 4.04 software (Social Information Service). One-way ANOVA was performed, and the mean differences among the three groups were evaluated using Tukey's methods with a significance level of 1%.

[0037] In particular, the shiitake mushroom extract has high glutaminase activity in the presence of sodium chloride, and therefore can increase the free glutamic acid content in the presence of sodium chloride, enhancing the umami taste of foods. In addition, the production of 5'-guanylic acid can have a synergistic effect in enhancing the umami taste.

[0038] (Enzyme purification and chromatographic analysis of Enokitake mushroom extract) As in the above-mentioned example, 2 kg of the edible portion of the Enokitake mushroom was mixed with 3 times the amount of 10 mM phosphate buffer (pH 7), homogenized, and centrifuged. The resulting supernatant was used as a crude extract and subjected to Westase treatment, followed by chromatography to fractionate the GAP activity peak.

[0039] For Westase (β1,3-glucanase) treatment, 80 mL of 0.5% Westase solution (pH 6.0) was added to 1.5 L of Enoki extract (crude extract) and incubated at 40°C for 30 minutes. TOYOPEARL DEAE 650M anion chromatography was performed using a 2.5 x 30 cm column at a flow rate of 1 mL / min with elution using a linear gradient of 0.1-0.6 M NaCl / 10 mM phosphate buffer (pH 7.0). TOYOPEAER L Butyl 650M hydrophobic chromatography was performed using a 2.5 x 30 cm column at a flow rate of 1 mL / min with elution using a linear gradient of 1.6-0 M ammonium sulfate / 10 mM phosphate buffer (pH 7.0). Ca++Pure-HA hydroxyapatite chromatography was performed using a 1.0 x 12 cm column with linear gradient elution using 5-200 mM sodium phosphate buffer (pH 7.0) at a flow rate of 1 mL / min. High performance liquid chromatography (HPLC) was performed using a gel filtration column TSKgel G3000SW at a column temperature of 40°C, a flow rate of 0.3 mL / min, and elution with 0.1 M NaCl / 10 mM phosphate buffer (pH 7.0).

[0040] (Native-Page and activity staining) Native PAGE (10% T gel) was performed at 20 mA for 90 minutes using an ATTO Pagelan AE-6531 electrophoresis tank. Activity staining was performed by immersing the gel after electrophoresis in a 5 mM Glu-pNA solution at 37°C for 30 minutes.

[0041] (Characteristics of GAP activity of partially purified enzyme) To determine the optimum pH, Britton-Robinson buffers ranging from pH 3 to pH 9 were used. pH stability was evaluated by mixing the partially purified enzyme with buffers ranging from pH 3.0 to 9.0, incubating at 37°C for 30 minutes, and then measuring the remaining GAP activity. For the optimum temperature, measurements were performed at pH 7.0 between 30 and 80° C. For thermal stability, the partially purified enzyme was kept at 30 to 80° C. for 30 minutes, and then the remaining GAP activity was measured at pH 7.0.

[0042] (Substrate specificity of partially purified enzyme) As a substrate, each amino acid pNA solution (2 mM, pH 7.0) was used, and the reaction was carried out at 37°C for 30 minutes. The increase in OD405 nm per minute was measured, and the amount of enzyme that produces 1 μmol of pNA per minute per mg of protein was defined as 1 U.

[0043] (Native and SDS-PAGE of gel filtration HPLC separated peaks) For native-PAGE, we used a homemade 10% T gel in an ATTO Pagelan AE-6531 electrophoresis tank, and for SDS-PAGE, we used a ready-made gel (T5-20% gradient gel).

[0044] (Effect of salt on GAP and PAP activities) GAP activity and PAP activity were measured using a Glu-pNA solution or a Pro-pNA solution containing 0 to 20% NaCl as a substrate, and the maximum activity of each was taken as 100%, and the relative activity was expressed.

[0045] Enokitake mushroom extract is expected to contribute to the production of free amino acids, and in particular, because it has high glutamyl aminopeptidase activity, it can liberate glutamic acid from glutamyl peptides. In addition, it can increase the free glutamic acid content in the presence of sodium chloride.

Claims

1. A food umami enhancer comprising an extract of shiitake mushroom or enoki mushroom, which is contacted with food in the presence of a chloride salt.

2. The umami enhancer according to claim 1 , wherein the contact is carried out in the presence of 0.1% to 20% of said chloride salt.

3. 2. The umami taste enhancer of claim 1, wherein the chloride salt is selected from the group consisting of sodium chloride, potassium chloride, magnesium chloride, ammonium chloride, and calcium chloride.

4. A method for enhancing the umami taste of a food product comprising contacting an extract of shiitake or enokitake mushroom with the food product in the presence of a chloride salt.

Images