Salty taste modifiers, and food and beverage products including salty taste modifiers

Abstract

Salty-taste modifier compositions are provided that include one or more salty taste-modifier peptides. The salty taste modifier compositions may be added to food or beverage products to modify, enhance, or add salty taste to the food or beverage product.

Description

SALTY TASTE MODIFIERS, AND FOOD AND BEVERAGE PRODUCTS INCLUDING SALTY TASTE MODIFIERSTechnical Field

[0001] This invention generally rela tes to salty taste modifiers and the inclusion of such salty taste modifiers in food and beverage products. More particularly, this disclosure generally relates to salty modifier peptides and the inclusion of such peptides in food and beverage products.Reference to a Sequence Listing[0002) The Sequence Listing associated with this application is filed in electronic form via Patent Center and is hereby incorporated by reference in its entirety. The name of the electronic file containing the Sequence Listing is '1410-163200-US_SL. The size of the.xml file is 21.1 KB and the file was created on December 17, 2025.Background{0003J According to the World Health Organization (WHO), a large number of people around the world are consuming too much sodium. The mean daily intake of sodium by ad ults globally is reportedly double the WHO's recommended intake of less than 2,000 mg / day.Efforts have been made by food companies to provide lower sodium alternatives, but those efforts have generally been met with reduced consumer acceptance of the food products.{0004J Improving consumer acceptance may require maintaining the flavor profile delivered by sodium chloride so that the reduced sodium product has the same flavor as a non¬ reduced sodium formulation. However, commercially available options for adding a salty taste to food products generally are characterized by a limited salty flavor impact and often deliver off flavors, including bitter or metallic flavors, that further limit consumer acceptance of the food products. For example, potassium chloride has been used as a salt substitute but it is often perceived as adding bitter flavor.

[0005] It has also been found that inclusion of sodium chloride in food products influences other aspects of the flavor of the food. It has been found that removal or reduction of salt can result in alteration of other flavors in the food. For example, perception of sweet, sour, and bitter flavors may increase or decrease when salt content is reduced in a food product.

[0006] Attempts have also been made to increase salty flavors in reduced sodium food products to help mitigate flavor changes. Commercially available salt-enhancing ingredients also often add other undesirable flavors, such as sour, bitter, or astringent flavors to the food or beverage products, which results in reduced consumer acceptance.Detailed Description

[0007] Described herein are salty taste modifier compositions that can be added to food or beverage products. The taste modifier compositions include one or more salty taste-modifier peptides that, either alone or in combination, are effective to modify salty taste to the food or beverage products, in some approaches, the salty taste modifier compositions modify, add, increase, and / or enhance a salty taste in the food or beverage product. Generally speaking, a salty taste is typically associated with the presence of salts, particularly sodium chloride (NaCl). It provides a sensation of savoriness and can enhance the overall flavor profile of a dish.Saltiness is often perceived as a basic taste that balances and complements other flavors. Foods such as chips, pretzels, and salted nuts are common sources of salty taste.

[0008] Salty is one of the five recognized main taste characteristics in food and beverage products, along with sweet, sour, bitter, and umami. In addition to those five characteristics, kokumi and astringency are also sometimes classified as taste characteristics. Generally speaking, sweetness is a taste typically associated with sugars, for example sucrose, glucose, and fructose, as well as artificial sweeteners. It is often perceived as pleasurable and is commonly found in desserts, fruits, and sugary beverages. Sourness is caused by acids, such as citric acid (found in citrus fruits), acetic acid (in vinegar), and lactic acid (in fermented dairy). The sour taste is sharp and tangy and can range from mildly tart to intensely acidic. It is a flavor commonly found in fruits like lemons, green apples, and in fermented products like yogurt or pickles. Acids can also act to improve the stability in some foods and beverages by lowering the pH of the system. Bitterness is a taste often associated with compounds like alkaloids (e.g.,Attorney Docket 1410-163200-PCcaffeine or quinine) or certain plant-based substances. It is usually perceived as unpleasant or sharp, and many bitter foods are avoided due to their strong flavor. Bitter tastes are found in foods such as dark chocolate, coffee, kale, and certain herbs. While it can be off-putting in large quantities, bitterness can also add complexity and balance to some dishes. Umami is often described as a savory or meaty taste, and it is produced by the presence of amino acids such as glutamate, as well as nucleotides like inosinate and guanylate. It is considered a rich, full- bodied flavor that enhances the taste of foods. Umami is commonly found in foods like meat, cheese, soy sauce, tomatoes, mushrooms, and fermented products. Kokumi is described as providing a background richness and complexity to foods, and particularly savory foods.Astringency is described as providing a puckering and drying feeling in the mouth.

[0009] As used herein, the term '' taste modifier" refers to a food ingredient that has the effect of modifying (either by increasing or decreasing) a perceived taste of at least one of the five main taste characteristics in food or beverage products when the flavor modifier is added to the food or beverage product. Also described herein are food and beverage compositions comprising the flavor modifier, as well as methods of making a food or beverage composition comprising the taste modifier.(0010) At least in some approaches, by modulating the perceived taste of the food or beverage product, one or more other ingredients may be reduced or removed from the food or beverage product without having a deleterious effect on the taste characteristic of the food or beverage product. In another approach, an existing taste characteristic may be increased in the food or beverage product by addition of the taste modifier.

[0011] In some approaches, the salty taste modifiers described herein are salty-enhancing taste modifiers. As used herein, the term "enhancing" means that the salty- enhancing modifiers add a salty taste or increase the intensity of a salty taste in the food or beverage product.

[0012] In one approach, the taste modifier is a salty-taste modifier. In some embodiments, the salty-taste modifiers described herein are added alone or in combination with one or more of the other salty taste modifiers to modify, add, enhance, and / or increase the intensity of salty taste to food or beverage products. The salty-taste modifiers may also be 3Attorney Docket 1410-163200-PCadded alone or in combination with one or more other taste modifying ingredients to food or beverage products.

[0013] For example, the addition of one or more of the salty -enhancing modifiers in an amount of at least about 0.0007 wt.%, at least about 0.0008 wt.%, at least about 0.0009 wt.%; at least about 0.001 wt.%, at least about 0.002 wt.%, at least about 0.003 wt.%, at least about 0.004 wt.%, at least about 0.0005 wt.%, at least about 0.006 wt.%, at least about 0.007 wt.%, at least about 0.008 wt.%, at least about 0.009 wt.%, at least about 0.01 wt.%, at least about 0.05 wt.%, at least about 0.1 wt.%, or at least about 1.0 wt% of the food or beverage product may allow for reduction of sodium.{0014J In one approach, by including one or more such salty-taste modifiers, in addition to enhancing the salty taste of a food or beverage product, the amount of sodium in the food or beverage product may be reduced. For example, the addition of one or more of the salty- enhancing modifiers may allow for reduction of sodium by at least about 10 percent, at least about 20 percent, at least about 25 percent, at least about 30 percent, at least about 35 percent, at least about 40 percent, at least about 45 percent, or at least about 50 percent.[0015| In another approach, the salty-enhancing modifier can be added in an amount effective to add salty taste to the food or beverage product but without adding off flavors, such as bitterness, that will reduce consumer acceptance of the food or beverage product. To achieve this, the salty-enhancing modifier has a salty modulation threshold concentration that is less than its intrinsic taste threshold concentration (e.g., bitter, astringent, or sour taste) in a given food or beverage product. In other words, the salty-enhancing effect of the salty-enhancing modifier detectable by taste in a given food or beverage product when the salty-enhancing modifier is added is achieved at a lower amount than the amount of the salty -enhancing modifier at which a bitter, astringent, or sour taste of the salty-enhancing modifier is detectable in the food or beverage product. Further, the lower the salty-enhancing threshold value, the more potent the sally-modifying effect in a food or beverage product, meaning that the salty -enhancing modifier may be included in a small quantity and still provide the desired salty-enhancing effect.Attorney Docket 1410-163200-PC

[0016] In one particular approach, the sally -enhancing peptides include one or more salty-enhancing peptides having the following amino acid sequences: EIV; HQPHQPLPP (SEQ ID NO. 8); and SKVLPVPQ (SEQ ID NO. 12). Any one of these salty-enhancing peptides may be added to food or beverage products alone, in combination with one or more of the other of these three salty-enhancing peptides, and / or in combination with other salty-enhancing peptides. When used in combination, the salty-enhancing peptides may be combined in any ratio to provide the desired flavor profile in the food or beverage product.|0017| In some aspects, the one or more salty -enhancing peptides are added to a food or beverage product as a taste modifying composition in liquid (e.g., as a solution or other aqueous mixture) or in powder form.

[0018] Tn some approaches, the one or more salty-enhancing peptides may be provided in a composition that includes other peptides, such as dairy-based peptides, but in which the one or more salty-enhancing peptides are included at a significantly higher amount than the other peptides. For example, the one or more salty-enhancing peptides may be individually included in an amount of at least 5 times, at least 10 times, at least 20 times, at least 30 times, at least 40 times, at least 50 times, at least 100 times, at least 500 times, or at least 1000 times the amount of other individual peptides in a composition.

[0019] In some approaches, the salty -enhancing peptides are added to the food and beverage products in substantially pure or isolated form. The salty-enhancing peptides may be isolated from other dairy-based proteins and / or peptides, such as other proteins and peptides found in dairy products like milk, cream, and natural cheese, isolated from other dairy-based compounds, or isolated from other peptides (such as other peptides found in dairy products). For example, each of the salty-enhancing peptides may be provided in isolated form, meaning that one of the salty-enhancing peptides is the primary peptide (e.g., at least 50 wt.% of the total peptides present) in a composition. Alternatively, a combination of the salty-enhancing peptides may be the primary peptide component (e.g., at least 50 wt.% of the total peptides present) in a composition. As used herein, the term "isolated" with respect to a salty-enhancing polypeptide composition means that the salty-enhancing polypeptide composition includes one or more of the salty-enhancing peptides in an amount of at least 60%, at least 70%, at least 80%, at leastAttorney Docket 1410-163200-PC90%, at least 95%, or at least 99% by dry weight of the salty -enhancing polypeptide compositions.

[0020] In certain implementations, the salty-enhancing peptides are added to a food or beverage product as a taste-modifier composition further comprising one or more carriers, which may also be a bulking agent, solvent, or other liquid. For example, the taste-modifier composition may be in a dry form such as a powder or particulate. In other approaches, the taste-modifier composition may be in liquid form, such as in the form of a fermentation extract or cultured milk. In yet other approaches, the taste-modifier composition may be in solid or semi-solid form, such as in the form of a flavor ingredient, concentrated cheese flavor, or enzyme-modified cheese. Suitable carriers include any food grade carrier that does not adversely impact the flavor of the food or beverage product. Exemplary food grade carriers include one or more of gum arabic, dextrose, sucrose, fructose, lactose, proteins, cellulose, silica, calcium silicate, tapioca flour, rice flour, salt, potassium chloride, starch, and / or maltodextrin, such as corn maltodextrin, potato maltodextrin, tapioca starch, modified tapioca starch, rice flour, and tapioca maltodextrin. For instance, the salty -enhancing peptides may be included in a taste-modifier composition in an amount of at least about 1%, at least about 2%, at least about 5%, at least about 10%, at least about 20%, by dry weight of the taste-modifier composition. In some approaches, the carrier may be the major component of the taste-modifier composition.

[0021] In some aspects, the one or more salty-enhancing peptides are added to a food or beverage product in an aqueous solution or mixture. In certain implementations, the salty-enhancing peptides, when added to a food or beverage product as a liquid taste-modifier composition, may be added together with one or more carriers including but not limited to water, glycerin, propylene glycol, 1,3-propanediol, ethanol, lactic acid, and combinations thereof.100221 In some aspects, the one or more salty -enhancing peptides are added to a food or beverage product in the form of a fermentation extract. For example, one or more of the salty - enhancing peptides may be developed within a specific product through targeted fermentation or enzymatic treatment aimed at obtaining a certain concentration. In some implementations, concentrated fermented or enzymatically produced "flavors" with high concentrations of thesalty-enhancing peptides may be generated from non-dairy sources or dairy sources. For example the one or more salty-enhancing peptides may be generated in the form of an enzyme- modified cheese, enzyme-modified fresh cheese, cultured milk, or the like, and then added as an ingredient into a food or beverage product. The one or more salty-enhancing peptides might also be added to a food or beverage product in the form of a flavor ingredient. The one or more salty-enhancing peptides may be generated from a dairy-based source in some approaches or a plant-based source in other approaches.

[0023] In one approach, the salty-enhancing peptides may be added to a food or beverage product to increase the salty taste of the food or beverage product. The food or beverage product may be any product that already has a salty taste or any product that might benefit from having an enhanced salty taste.

[0024] In certain aspects, the salty-enhancing peptides may be added to a food or beverage product in which it is desired to reduce the sodium content thereof. At least in some approaches, the addition of the salty-enhancing peptides adds salty taste to the food or beverage product, which may counteract, ameliorate, or otherwise mitigate a reduction of the sodium amount in the food or beverage product.

[0025] In some aspects, the amount of salt in the food or beverage product can be reduced as a result of the addition of the one or more salty-enhancing peptides to the food or beverage in sufficient amounts. In some aspects, inclusion of one or more of the salty-enhancing peptides may allow for reduction of sodium by at least about 10%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, or at least about 50%. In some aspects, the amount of salt in the food or beverage product may be reduced by about 10% to about 50%, by about 15% to about 45%, by about 20% to about 40%, by about 25% to about 35%, or by about 30%, as compared to a substantially identical food or beverage product to which the one or more salty -enhancing peptides have not been added (with the weight percentages of the ingredients of this substantially identical food or beverage product being identical to the food or beverage product containing the salty-enhancing peptides, except for water, bulking agent (e.g., a major bulk contributor in a given food product, such as tomato paste in ketchup), or the like, which is included in the substantially identical7Attorney Docket 1410-163200-PCfood products in an amount sufficient to replace the one or more salty-enhancing peptides present in the food or beverage product containing the salty -enhancing peptides).

[0026] Suitable food products to which the salty-enhancing peptides described herein may be added include but are not limited to: process cheese (e.g., in slice, loaf, or liquid sauce form), cream cheese (e.g., dairy cream cheese, plant-based cream cheese, flavored cream cheese (e.g., chive & onion, jalapeno, etc,), unflavored cream cheese (e.g., plain, etc.)), plant -based cheese, shredded cheese, natural cheese, dry meals (e.g., macaroni and cheese powder, rice (e.g., rice mix, etc.), stuffing mix, pasta (e.g., pasta mix, macaroni and cheese), etc.), sauces (e.g., cheese sauce, tomato frito, cocktail sauce, sweet and sour sauce, barbeque sauce, soy sauce, tartar sauce, hot sauce, steak sauce, pasta sauce (e.g., tomato sauce, alfredo sauce, etc.)), and condiments (e.g., tomato ketchup, mustard, mayonnaise, aioli, salad dressing, dairy or non¬ dairy creamy dressings (e.g., ranch dressing, green goddess, Caeser, etc.), oil-based dressings (e.g., vinaigrette, Italian dressing, French, etc.), gravy (ready-to-use or dry), frozen meals and / or sides (e.g., frozen potato products such as French fries, potato tots, beans, etc.), soups (e.g., chicken broth-based soup), beans, or other meals, and dry seasonings (e.g., taco seasoning, etc.) and foods containing such dry seasonings (e.g., potato chips, crackers, etc.). All of the food products mentioned herein refer to either a dairy-based or plant-based version of the food product unless specifically stated otherwise. In some approaches, the food or beverage product to which the one or more salty -enhancing peptides are added are not dairy products, in some approaches, the food or beverage product to which the one or more salty -enhancing peptides are added are not cheese products.[00271 In some approaches, the food products are dairy-based products. As used herein, the term “ daily -based" refers to a product or ingredient that contains one or more ingredients originating from dairy milk or cream. For example, dairy-originating ingredients include whey protein, milkfat, cream, milk, milk solids (e.g., non-fat dry milk), butter, milk-based yogurt, cheese curd, cheese, and the like, as well as food ingredients that include non-animal based dairy proteins, such as whey or fermented whey proteins that may be obtained from non¬ animal sources.8Attorney Docket 1410-163200-PC

[0028] In some approaches, the food products are plant- based products. As used herein, the term "plant-based" refers to a product or ingredient that is free of animal-based ingredients, such as proteins (e.g., dairy proteins like casein and whey) and fats (such as milk fats). The plant-based products may instead include a plant-based protein. Any suitable plant-based protein may be used in the plant-based product. In some aspects, the plant-based protein comprises one or more of faba bean protein (also known as fava bean protein), soy protein, lentil protein, potato protein, chickpea protein, canola protein, and pea protein. It has been found that some plant-based proteins may impart an off flavor to the resulting plant-based product. Therefore, inclusion of one or more salty -modifier peptides may be particularly beneficial for plant-based food products.{0029J In one approach, the salty-enhancing peptides include one or more of the following peptides: EIV peptide; HQPHQPLPP (SEQ ID NO. 8) peptide; and SKVLPVPQ (SEQ ID NO. 12) peptide. Advantageously, each of EIV peptide, HQPHQPLPP (SEQ ID NO. 8) peptide, and SKVLPVPQ (SEQ ID NO. 12) peptide has a salty modulating threshold concentration in a model broth (including sodium chloride, monosodium L-glutamate, maltodextrin, yeast extract, and water) that is lower than its intrinsic taste threshold concentration, which means that these three salty-enhancing peptides can be added to a food product in an amount that provides salty-enhancing effect without also adding a bitter or sour flavor to the food product. This characteristic makes these three peptides particularly advantageous as a salty-enhancing ingredient.

[0030] To enhance the salty taste of a given food or beverage product, any one of the following seven salty taste enhancing compositions may he added to the food or beverage product: (1) EIV peptide; (2) HQPHQPLPP (SEQ ID NO. 8) peptide: (3) SKVLFVPQ (SEQ ID NO. 12) peptide: (4) EIV peptide and HQPHQPLPP (SEQ ID NO. 8) peptide: (5) EIV and SKVLFVPQ (SEQ ID NO. 12) peptide: (6) HQPHQPLPP (SEQ ID NO. 8) peptide and SKVLPVPQ (SEQ ID NO. 12) peptide; (7) ETV peptide, HQPHQPLPP (SEQ ID NO. 8) peptide, and SKVLPVPQ (SEQ ID NO. 1 ) peptide. It will be appreciated that any one of these seven salty-enhancing compositions may further include one or more other taste-modifier (e.g., a salty-enhancing, bitter-enhancing. umami-enhancing, kokumi-enhancing, sweet-enhancing, and / or astringent-enhancing) peptides and / or salty-enhancing compounds known in the art.9Attorney Docket 1410-163200-PC

[0031] The salty taste enhancing compositions may include any amount of the peptide(s), alone or in combination, necessary to provide a saity-enhancing effect when the salty taste enhancing composition is added to a food or beverage product. For example, the salty taste enhancing compositions may include one or more of the salt -enhancing peptides in an amount, by dry weight, of at least 0.1 wt.%, at least 0.5 wt.%, at least 1 wt.%, at least 2 wt.%, at least 5 wt.%, at least 10 wt.%, at least 15 wt.%, at least 20 wt.%, at least 25 wt.%, at least 30 wt.%, at least 35 wt.%, at least 40 wt.%, at least 45 wt.%, at least 50 wt.%, at least 60 wt.%, at least 70 wt.%, at least 80 wt.%, or at least 90 wt.% of the taste modifier composition. In another approach, the salty taste enhancing compositions may include one or more of the salty-enhancing peptides in an amount, by total weight, of at least 0.1 wt.%, at least 0.5 wt.%, at least 1 wt.%, at least 2 wt.%, at least 5 wt.%, at least 10 wt.%, at least 15 wt.%, at least 20 wt.%, at least 25 wt.%, at least 30 wt.%, at least 35 wt.%, at least 40 wt.%, at least 45 wt.%, at least 50 wt.%, at least 60 wt.%, at least 70 wt.%, at least 80 wt.%, or at least 90 wt.% of the taste modifier composition. The salty taste enhancing composition may then be added in any desired dilution factor or ratio to provide the food or beverage product with a salty-enhancing amount of one or more of the peptides.

[0032] Table 1 provides the salty enhancing taste threshold for each peptide (i.e., the minimum concentration to modulate salty taste) in model broth. The salty enhancing taste threshold in model broth for each peptide may provide a starting point when formulating a food or beverage product containing one or more of the salty enhancing peptides. However, it has been found that the salty enhancing taste threshold for one or more of the peptides is lower in certain food and beverage products than in the model broth. Therefore, the amount of one or more of the peptides may be lower than the salty enhancing threshold provided in Table 1. For those food and beverage products, an amount of 0.25x salty enhancing threshold is provided as a proposed minimum amount for each of the salty enhancing peptides.Table 1: Taste Thresholds for Salty-Enhancing PeptidesSalty Taste Enhancing Peptides | Amount Present in Cheddar Cheese in mg / kg (wt.%) | Salty Taste Enhancing Threshold in Model Broth | Intrinsic Taste Threshold in Water | 0.25x Salty Taste Enhancing Threshold in Model BrothEIV 0.05 mg / kg 7.4 mg / L 20.3 mg / L 0.000175 wt.%(bitter) i(0.000005 wt.%) (0.0007wt. %) (0.00203%)HQPHQPLPP (SEQ ID 7.02 mg / kg 18.2 mg / L 125.8 mg / L | 0.00045 wt.% NO. 8) (sour) i(0.000702 wt.%) (0.0018wt.%) (0.01258 wt.%)SKVLPVPQ (SEQ ID 6.30 mg / kg 41.3 mg / L 82.7 mg / L 0.001025 wt.% NO. 12) (bitter) |(0.00063 wt.%<) (0.0041wt.%) (0.00827 wt.%) |

[0033] In some embodiments, one or more of the above-mentioned salty taste- enhancing compositions may be added to a food or beverage product in an amount effective to provide a salty taste enhancing effect. The salty taste-enhancing peptides may be added alone or in any combination in an amount effective to provide a desired salty enhancing taste to a food or beverage product. In some approaches, the salty taste-enhancing peptides are included in an amount lower than an amount that provides an undesirable taste (e.g., below an amount that provides an intrinsic taste of bitter or sour). The salty taste enhancing thresholds identified in Table 1 are based on inclusion of the peptides in model broth. However, amounts lower than the salty taste enhancing threshold in model broth may still provide a desirable salty tasteenhancing effect in other food or beverages. Also, in some food or beverages, an undesirable intrinsic taste may be partially or completely masked by other ingredients in the product, Additionally, off-flavor masking ingredients (e.g., bitter blocking or masking ingredients) may be added to the food or beverage products. Therefore, it is contemplated that each of the salty- enhancing peptides may be added in higher amounts than their intrinsic taste threshold.

[0034] In some aspects, a food or beverage composition includes at least about 0.0007 wt.%, at least about 0.0008 wt.%, at least about 0.0009 wt.%, at least about 0.001 wt.%, at least about 0.005 wt.%, at least about 0.01 wt.%, at least about 0.05 wt.%, at least about 0.1 wt.%, at least11Attorney Docket 1410-163200-PCabout 0.5 wt.%, or at least about 1.0 wt.% of the EIV peptide to provide a desired salty enhancing effect. In some approaches, the EIV peptide may be included in a food or beverage product in an amount of at least its taste threshold value (i.e., 0.0007 wt.%) model broth but below its intrinsic taste value in water (i.e., 0.00203 wt.% with a bitter taste). For example, a food or beverage composition may include about 0.0007 wt.% to less than 0.00203 wt.%, about 0.0008 wt.% to less than 0.00203 wt.%, about 0.0009 wt.% to less than 0.00203 wt%, about 0.001 wt.% to less than 0.00203 wt.%, or about 0.0015 wt.% to less than 0.00203 wt%, of the EIV peptide to provide a salty enhancing effect.

[0035] In other food or beverage systems, the EIV peptide may be effective to provide a salty enhancing effect at a lower concentration than its salty taste threshold in model broth. For example, the EIV peptide may be included in a food or beverage product In an amount of at least 0.25x its taste threshold in model broth. The EIV peptide may be included in a food or beverage product in an amount of at least 0.000175 wt.%, at least 0.0002 wt.%, at least 0.0003 wt.%, at least 0.0004 wt.%, at least 0.0005 wt.%, or at least 0.0006 wt.%. In another approach, the EIV peptide may be included in a food or beverage product in an amount of at least 0.000175 wt.% to less than 0.00203 wt.%, at least 0.0002 wt.% to less than 0.00203 wt.%, at least 0.0003 wt.% to less than 0.00203 wt.%, at least 0.0004 wt.% to less than 0.00203 wt.%, at least 0.0005 wt.% to less than 0.00203 wt.%, at least 0.0006 wt.% to less than 0.00203 wt.%, at least 0.0007 wt.% to less than 0.00203 wt.%, at least 0.0008 wt.% to less than 0.00203 wt.%, at least 0.0009 wt.% to less than 0.00203 wt.%, or at least 0.001 wt.% to less than 0.00203 wt.% to provide a salty enhancing effect.[0036) In some aspects, a food or beverage composition includes at least about 0.0018 wt%, at least about 0.002 wt.%, at least about 0.003 wt.%, at least about 0.004 wt.%, at least about 0.005 wt%, at least about 0.006 wt.%, at least about 0.007 wt.%, at least about 0.008 wt.%, at least about 0.009 wt.%, at least about 0.01 wt%, st least about 0.02 wt.%, at least about 0.03 wt.%, at least about 0.04 wt.%, at least about 0.05 wt%, at least about 0.1 wt%, or at least about 1.0 wt% of the HQPHQPLPF peptide (SEQ ID NO. 8) to provide a salty enhancing effect. In some approaches, the HQPHQPLPP peptide (SEQ ID NO. 8) may be included in a food or beverage product in an amount of at least its taste threshold value (i.e., 0.0018 wt.%) in model broth but below its intrinsic taste value in water (i.e., 0.01258 wt.%). For example, a food or beverageAttorney Docket 1410-163200-PCcomposition may include about 0.0018 wt.% to less than 0.01258 wt.%, about 0.002 wt.% to less than 0.01258 wt.%, about 0.003 wt.% to less than 0.01258 wt.%, about 0.004 wt.% to less than 0.01258 wt.%, about 0.005 wt.% to less than 0.01258 wt.%, about 0.006 wt.% to less than 0.01258 wt.%, about 0.007 wt.% to less than 0.01258 wt.%, about 0.008 wt.% to less than 0.01258 wt.%, about 0.009 wt.% to less than 0.01258 wt.%, or about 0.01 wt.% to less than 0.01258 wt.% of the HQPHQPLPP peptide (SEQ ID NO. 8) to provide a salty enhancing effect.

[0037] In other food or beverage systems, the HQPHQPLPP peptide (SEQ ID NO. 8) may be effective to provide a salty enhancing effect at a lower concentration than its salty taste threshold in model broth. For example, the HQPHQPLPP peptide (SEQ ID NO. 8) may be included in a food or beverage product in an amount of at least 0.25x its taste threshold in model broth. The HQPHQPLPP peptide (SEQ ID NO. 8) may be included in a food or beverage product in an amount of at least 0.00045 wt.%, at least 0.0005 wt.%, at least 0.0006 wt.%, at least 0.0007 wt.%, at least 0.0008 wt.%, at least 0.0009 wt.%, or at least 0.001 wt.% to provide a salty enhancing effect. In another approach, the HQPHQPLPP peptide (SEQ ID NO. 8) may be included in a food or beverage product in an amount of at least 0.00045 wt.% to less than 0.01258 wt.%, at least 0.0005 wt.% to less than 0.01258 wt.%, at least 0.0006 wt.% to less than 0.01258 w't.%, at least 0.0007 wt.% to less than 0.01258 wt.%, at least 0.0008 wt.% to less than 0.01258 wt.%, at least 0.0009 wt.% to less than 0.01258 wt.%, at least 0.001 wt.% to less than 0.01258 wt.%, at least 0.002 wt.% to less than 0.01258 wt.%, at least 0.003 wt.% to less than 0.01258 wt.%, at least 0.004 wt.% to less than 0.01258 wt.%, at least 0.005 wt.% to less than 0.01258 wt.%, at least 0.006 wt.% to less than 0.01258 wt.%, at least 0.007 wt.% to less than 0.01258 wt.%, at least 0.008 wt.% to less than 0.01258 wt.%, at least 0.009 wt.% to less than 0.01258 wt.%, or at least 0.01 wt.% to less than 0.01258 wt.%, to provide a salty enhancing effect.

[0038] In some aspects, a food or beverage composition includes at least about 0.0041 wt%, at least about 0.005 wt.%, at least about 0.006 wt.%, at least about 0.007 wt.%, at least about 0.008 wt.%, at least about L’. OL’y wt.%, at least about 0.01 wt%, at least about 0.0t> wt%, at least about 0.1 wt%, at least about 0.5 wt%, or at least about 1.0 wt% of the SKVLPVPQ peptide (SEQ ID NO. 12) to provide a desired salty enhancing effect. In some approaches, the SKVLPVPQ peptide (SEQ ID NO. 12) may be included in a food or beverage product in an amount of at least its taste threshold value (i.e., 0.0041 wt.%) in model broth but below its intrinsic taste value 13Attorney Docket 1410-163200-PCin water (i.e., 0.00827 wt.%). For example, a food or beverage composition may include about 0.0041 wt.% to less than 0.00827 wt.%, about 0.005 wt.% to less than 0.00827 wt.%, about 0.006 wt.% to less than 0.00827 wt.%, about 0.007 wt.% to less than 0.00827 wt. %, or about 0.008 wt. % to less than 0.00827 wt.% of the SKVLPVPQ peptide (SEQ ID NO. 12) to provide a salty enhancing effect.

[0039] In other food or beverage systems, the SKVLPVPQ peptide (SEQ ID NO.12) may be effective to provide a salty enhancing effect at a lower concentration than its salty taste threshold in model broth. For example, the SKVLPVPQ peptide (SEQ ID NO. 12) may be included in a food or beverage product in an amount of at least 0.25x its taste threshold in model broth. The SKVLPVPQ peptide (SEQ ID NO. 12) may be included in a food or beverage product in an amount of at least 0.001025 wt.%, at least 0.002 wt.%, at least 0.003 wt.%, at least 0.004 wt.%, at least 0.005 wt.%, at least 0.006 wt.%, at least 0.007 wt.%, or at least about 0.008 wt.% to provide a salty enhancing effect. In another approach, the SKVLPVPQ peptide (SEQ ID NO. 12) may be included in a food or beverage product in an amount of at least about 0.001025 wt.% to less than 0.00827 wt.%, about 0.002 wt.% to less than 0.00827 wt.%, about 0.003 wt.% to less than 0.00827 wt.%, about 0.004 wt.% to less than 0.00827 wt.%, about 0.005 wt.% to less than 0.00827 wt.%, about 0.006 wt.% to less than 0.00827 wt.%, about 0.007 wt.% to less than 0.00827 wt.%, or about 0.008 wt.% to less than 0.00827 wt.%, of the SKVLPVPQ peptide (SEQ ID NO.12) to provide a sally enhancing effect.

[0040] Without wishing to be limited to theory, the salty-enhancing peptides can be produced via any suitable means in the art. In one approach, the salty -enhancing peptides may be derived or isolated from one or more cheddar cheeses, such as using an extraction method, in some aspects, the salty-enhancing peptides can be produced using enzymatic treatments for casein proteolysis and selecting enzymes that produce the desired salty-enhancing peptides in high yields.

[0041] For example, chemical synthesis may be used, such as solid phase peptide synthesis (SPPS), which is a well-known synthesis procedure. SPPS allows for assembly of a peptide chain by carrying out successive reactions of amino acid derivatives on a solid support. The Fmoc / fBu solid phase peptide synthesis may also be used. In some aspects, LPFS (liquid14Attorney Docket 1410-163200-PCphase peptide synthesis), fragment condensation, native ligation or recombinant methods may be used for peptide synthesis.

[0042] In some aspects, when SPPS is used, the peptide assembly is conducted from the C to the N terminus, where the initial ammo acid or short-chained peptide is covalently linked to a solid support material (e.g., resin). Next, a solution of the desired amino acid with a protecting group at the N terminus and a second protecting group at the side chain is reacted with the previous molecule in a coupling reaction. It is added in excess and washed away after the reaction. The solid support will ensure easy reagent removal, and the protective groups will ensure selective reaction. After this, the N terminus of the newly added amino acid is deprotected (typically under basic or acidic conditions, depending on the protective group), and the next arnino acid can be added. This is repeated until the desired peptide has been assembled. Finally, the peptide is cleaved from the solid support and the amino acid side chains are deprotected to yield the final peptide product. For high peptide purities (as desired for use in food and beverages), the peptides will likely need to be purified (e.g., by RP-HPLC) to remove any traces of protective groups, reagents, or shorter-chained peptides.J0043] In some aspects, the salty-enhancing peptides described herein may be generated via recombinant techniques. For example, recombinant microorganisms (e.g., known bacterial cultures used in cheddar cheese production) comprising one or more genes whose expression results in synthesis of one or more of the salty-enhancing peptides may be used. In some approaches, the genes used in recombinant peptide / peptide precursor synthesis may be designed,{0044] In some aspects, lactic acid bacterial proteases may be used as recombinant enzymes that could generate one or more of the salty-enhancing peptides directly from casein. In one aspect, the EIV peptide can be formed from αS1-casein or β-casein, and HQPHQPLPP (SEQ ID NO. 8) and SKVLPVPQ (SEQ ID NO. 12) peptides can be formed from β-casein. In some aspects, the salty-enhancing peptides may be generated through fermentation and aging of cheddar cheese (but may likewise be generated through fermentation and aging of other cheeses). Without wishing to be limited to theory, the longer natural cheeses have been aged, the greater concentration of the peptides would be expected to be present in such cheeses. InAttorney Docket 1410-163200-PCcertain aspects, expression of the one or more genes whose expression results in synthesis of one or more of the salty-enhancing peptides may be inducible (i.e., activated under specific conditions or in the presence of specific compounds, providing control over when the gene is expressed in the recombinant microorganism).[00451 Also contemplated herein are genes encoding a polypeptide comprising the amino acid sequence of the EIV peptide. Also contemplated herein are genes encoding a polypeptide comprising the amino acid sequence of SEQ ID NO. 8. Also contemplated herein are genes encoding a polypeptide comprising the amino acid sequence of SEQ ID NO. 12.[0046) Also contemplated herein are genes encoding a polypeptide having at least 80% identity, at least 90% identity, at least 95% identity, at least 99 % identity, or 100% identity to the amino acid sequence of the EIV peptide.. Also contemplated herein are genes encoding a polypeptide having at least 80% identity, at least 90% identity, at least 95% identity, at least 99% identity, or 100% identity to the amino acid sequence of SEQ ID NO. 8. Also contemplated herein are genes encoding a polypeptide having at least 80% identity, at least 90% identity, at least 95% identity, at least 99% identity, or 100% identity to the amino acid sequence of SEQ ID NO. 12.

[0047] In some approaches, the peptides may include additional amino acids than in the EIV peptide, the HQPHQPLPP (SEQ ID NO. 8) peptide, and the SKVLPVPQ (SEQ ID NO. 12) peptide so long as the additional amino acids do not significantly adversely impact rhe salty taste enhancing effect of the peptides.

[0048] In certain aspects, one or more of the genes for the salty-enhancing peptides described herein may be expressed in a recombinant host to produce one or more of the salty- enhancing peptides in a reproduceable and efficient manner. The recombinant host may be grown in any suitable media effective to result in production of one or more of the salty - enhancing peptides. For example, a single recombinant host may be used to generate the salty -enhancing EIV peptide, the salty-enhancing peptide of SEQ NO. 8, and the salty -enhancing peptides of SEQ ID NO. 12. The recombinant host may also be used to generate other salty - modifying or other taste-modifying peptides. The recombinant host may also be used to generate one or more aroma compounds of interest.16Attorney Docket 1410-163200-PC

[0049] In another approach, multiple recombinant hosts may be used to generate the salty-enhancing peptides. For example, a first recombinant host may be used to generate the salty-enhancing EIV peptide, a second recombinant host may be used to generate the salty- enhancing peptides of SEQ NO. 8, and a third recombinant host may be used to generate the salty-enhancing peptides of SEQ ID NO. 12. These recombinant hosts may be cultured separately or in mixed culture to generate the respective salty-enhancing peptides. Additional recombinant hosts may also be used to generate one or more aroma compounds of interest.100501 Possible recombinant hosts that may be suitable for this purpose include, but are not limited to Escherichia coli, Pichia pastoris, Lactococcus lactis, Bacillus subtilis, Saccharomyces cerevisiae, and mammalian cell lines. In some aspects, a suitable recombinant host is E. coli, which can be grown in Terrific broth culture media at 37°C for about 36 hours. Any suitable recombinant host or culture media may be used. After the culturing step, the salty -enhancing peptides thus produced can be recovered from the culture media using any suitable technique in the art. The salty-enhancing peptides may be concentrated or otherwise purified from the culture media by, for example, Ni-NTA affinity chromatography. At least in some approaches, the salty-enhancing peptides are recovered in substantially pure form, meaning that the one or more salty -enhancing peptides are the primary ingredient recovered from the culture media.

[0051] In some approaches, the recombinant host(s) may be used to produce a cultured milk containing one or more of the salty-enhancing peptides. For example, the recombinant host may be cultured in milk, concentrated milk, or other dairy liquid. In yet other approaches, the milk may be a plant-based milk, such as soy, oat, almond, cashew, chickpea protein, coconut, macadamia nut, pea protein, fava protein, or other aqueous liquid containing a plant protein source.100521 The following example is intended to illustrate the production and use of the salty-enhancing peptides and not to limit or otherwise restrict the disclosure. Unless indicated otherwise, all parts, ratios, and percentages are based on weight.

[0053] EXAMPLESAttorney Docket 1410-163200-PC

[0055] Extraction and Isolation of Taste- Active Compounds from Cheddar Cheese

[0056] Sensoproteomics was used to identify candidate taste-modifier peptides from cheddar cheese. As generally described in literature by Sebald et al. (2018), sensoproteomics combines sensory analysis (the study of how humans perceive taste, aroma, and texture) with proteomics (the study of proteins, particularly their functions and interactions). As such, sensoproteomics involves analyzing peptide sequences that may be responsible for influencing the taste of a product.. As a result of the procedure followed in this example, twelve salty- enhancing peptides were identified, and three of those salty-enhancing peptides were found to have particularly favorable salty-enhancing taste thresholds.

[0057] Activity-guided fractionation was performed on cheddar cheese to identify taste active peptide fractions. The cheddar cheese selected for the experiment had a strong cheddar flavor. Activity-guided fractionation is a technique used in natural product research to isolate and identify bioactive compounds from complex mixtures with the goal of identifying the specific compounds responsible for a particular activity (in this case, taste) by fractionating the mixhire and testing each fraction for its activity.

[0058] The activity -guided fractionation involved extracting peptides from cheddar cheese to obtain a crude extract. For extraction, 80 g of cheddar cheese was homogenized with 300 ruL 0.1% (v / v) formic acid in water (MilliQ Reference system with Elix 3, Millpore, Molsheim, France) using an Ultra-Turrax T 25 digital ilka Labortechnik, Staufen, Germany) at 13000 rpm for 5 min. The mixture was centrifuged (Avanti J-E Centrifuge, Beckmann Co lter, Brea California, US) at 4 °C and 10000 rpm for 30 min. The supernatant was collected, and the extraction residue was extracted twice more with 300 mb 0.1% (v / v) formic acid, respectively. The combined supernatants were lyophilized to yield the crude extract, which was stored at -20 °C until further use.

[0059] The crude extract was then subjected to fractionation using Medium Pressure Liquid Chromatography (MPLC) to separate the crude extract into smaller, more manageable fractions, each containing a different combination of compounds.

[0060] The MPLC instrument had two detectors, an evaporative light scattering detector (ELSD) and a U V detector. The UV detector was set to 220 nm based on preliminary experiments to optimize signal intensity. While UV detection is limited to UV-active compounds, ELSD is activated by most compounds present in a sample, including the investigated peptides. The detector signals indicate which fractions contain the highest amount of compounds. The information from the ELSD was used to determine how many fractions to pool for sensory analysis and to determine which fractions contain high concentrations of potentially taste-active compounds. However, the most active fractions were selected based only on TDA and cTDA, from which the TD- and cTD-factors were obtained. TD and cTD factors were indicative of the fraction(s) with the strongest taste impression and the strongest taste-modulating activity, respectively (i.e., the highest TD factor corresponds to the most taste¬ active fraction and the cTD factor corresponds to the most taste-modulating fraction).

[0061] The crude extract (3 g) was re-dissolved in 0.1% (v / v) formic acid in water (10 ml.) and fractionated using MPLC (Biichi, Flawil Switzerland), equipped with a reversed phase (RP) polypropylene cartridge filled with Chromabond Flash RS 120 C18 ec bulk material (40- 63 urn, 60 A, 26.2 cm x 37.2 mm, Macherey-Nagel, Dueren, Germany) with an injection volume of 2 mL, Separation was performed at a 40 mL / min flow rate using 0.1 % formic acid in water (A) and methanol (B) as solvents. After injection at 0% B, the concentration was held constant for 5 min, before increasing to 100% B within 50 min, and another isocratic step of 15 min. The concentration wras then decreased to 0% B within 12 min and held constant for 8 min.

[0062] Fractions F1–F60 were collected in one-minute intervals, and aliquots (1 mL) were dried under nitrogen, dissolved in 100 pL acetonitrile / water (50 / 50, v / v), and used for Liquid Chromatography / Time-of-Flight / Mass Spectrometry (" LC-ToF-MS") analysis followed by MaxQuant evaluation, as described in more detail below. The remainder of the MPLC fractions were pooled for fractions over 3 min each (fraction 1-3, 4-6, 7-9, 10-12, 13-15, 16-18, 19-21, 22-24, 25-27, 28-30, 31-33, 34-36, 37-39, 40-42, 43-45, 46-48, 49-51, 52-54, 55-57, and 58-60), freed from solvent in vacuo, and lyophilized twice to ensure complete solvent removal before sensory analysis via taste dilution analysis (TDA) and comparative taste dilution analysis (cTDA). The dried powders were stored at -20 °C until further use.

[0063] The most active (i.e., more-taste-active) fractions were then evaluated by a trained human sensory panel for flavor characteristics by Taste Dilution Analysis (TDA).Generally, TDA is a sensory technique used to evaluate the strength and detection threshold of specific flavors or taste compounds in food or beverages. TDA was used as a screening tool to identify the most taste-active MPLC fractions. It is typically not done for single compounds but rather compound mixtures such as the MPLC fractions to narrow down the number of compounds that could be responsible for the taste of the food. When looking for salty taste¬ active compounds, the focus was on the MPLC fractions that have the most intense salty taste m the TDA. After identification of the taste-active fractions, further purification can be used to isolate the specific compound(s) (e.g., peptides) responsible for this activity.

[0064] TDA was performed both in water (to determine "intrinsic taste") and in a model broth system (to determine "taste-modulating activity", cTDA). Duo-trio tests in increasing concentrations were used to determine the taste dilution (TD) factors for each fraction. A duo - trio test is a sensory test that determines if there is a difference between two samples by presenting a reference sample and two coded samples. The tasting panel was presented with a reference sample and then two coded samples, one of which matched the reference sample. The tasting panel then identified the coded sample that was different from the reference sample.

[0065] The panelists were asked to taste each fraction in several dilution steps and the taste dilution (TD) factor was calculated, which is the dilution factor at which the panelists can only just taste a difference between the sample and a blank (water). In this test, four panelists were used, and the TD-factors represented an average of the four individual TD-factors. A high TD factor is representative of the fact that this specific fraction is particularly taste-active. A fter the TD factors were determined, the most taste-active fractions (i.e., the fractions with the highest TD factors) were identified.

[0066] For TDA in water, each fraction was separately diluted in water beginning at natural cheese concentration and then diluted 1:2 in stepwise fashion to a total of 15 dilution steps. The natural cheese concentration was calculated from the amount of crude extract which was separated via MPLC. Since at this point it was not known which compounds were presentin the fractions and in which concentrations, this was only a qualitative comparison between the MPLC fractions to find out which fractions contain the taste-active compounds of interest.

[0067] The panelists started with the lowest concentrations. The panelists rinsed their mouths with water before each sample and wore a nose clip for all taste sensory tests to exclude cross-modal interactions by odor-active compounds. The results are presented in Table 2. Table 2 includes the most taste-active fractions that were identified. Table 3 below represents the full set of TD-factors for all of the tested fractions, from which the fractions shown in Table 2 were picked out as the most taste-active fractions. In Table 3, the underlined TD-factors represent the TD-factors of the fractions selected as the most taste-active fractions for Table 2.

[0068] Table 2 - TDA of Most Taste-Active MPLC Fractions in WaterFraction Number of Postulated Taste QualityPeptides4-6 1 Salt, umami, sour7-9 2 Sour13-15 7 Sour, salty28-30 143 A stringent, u ma mi31-33 231 Astringent, bitter

[0069] Table 3 - TDA of all Tested MPLC Fractions in Water and cTD A in Model Broth Fraction TD- Taste quality in cTD- TasteNumber of factor in water factor in modulating p o stul a ted p ep ti d es water model activity in based on MaxQuant broth model broth (score > 50)1-3 2 bitter 4 unspecific taste 04-6 431 sally, umami, 27 salty, umami, 1sour sweet7-9 215 sour 27 sour, salty 210-12 10 sour, astringent 16 umami 513-15 23 sour, salty 10 salty, sour 716-18 2 unspecific taste unspecific taste 1421Attorney Docket 1410-163200-PC19-21 2 unspecific taste 7 umami 822-24 5 astringent 7 salty 4025-27 4 astringent 3 salty 5928-30 32 astringent, umami 6 salty 14331-33 13 astringent, bitter 6 umami 23134-36 2 astringent 8 salty 9837-39 3 astringent 8 unspecific taste 9740-42 8 astringent, 32 umami, sweet 223burning43-45 5 bitter 32 umami 9146-48 5 bitter, astringent 8 unspecific taste 7049-51 2 astringent 8 salty 5052-54 10 sour, salty 32 salty 21255-57 8 unspecific taste 5 unspecific taste 1058-60 3 astringent 16 unspecific taste 0

[0070] For cTDA (comparative taste-dilution analysis) in the model broth system, a model broth as prepared by mixing 29 g NaCl, 1.9 g monosodium L-glutamate, 6.4 g maltodextrin, and 2.1 g yeast extract in water (1 L). Similarly to TD A, each fraction was separately diluted in model broth beginning at natural cheese concentration as a starting concentration and then diluted 1:2 in stepwise fashion to a total of 12 dilution steps. Sensory evaluation was conducted as detailed for TDA, and the comparative taste dilution (cTD)-factors were calculated for each fraction to identify the most active fractions (i.e. highest taste- modula ting activity).

[0071] The results of TD A revealed that fractions 4-6 (TD factor ~ 430), 7-9 (TD factor ~ 215), 13-15 (TD factor == 23), 28-30 (TD factor = 32) and 31-33 (TD factor - 13) had the highest intrinsic taste activity, whereas fraction 4-6 was perceived as salty, umami, and sour, fraction 7- 9 was sour, fraction 13-15 was sour and salty, fraction 28-30 was astringent and umami, and fraction 31-33 was astringent and bitter. In addition, cTDA revealed that fractions 4-6, 7-9, 40- 42, 43-45, and 52-54 were the most taste-modulating for salty, umami and sweet (fraction 4-6),22Attorney Docket 1410-163200-PCsour and salty (fraction 7- 9), umami and sweet (fraction 40–42), salty and umami (fraction 43–45) and salty (fraction 52–54) taste.

[0072] The fractions were further investigated using LC-ToF-MS analysis and MaxQuant evaluation, which revealed the presence of 1 (fraction 4-6), 2 (fraction 7-9), 7 (fraction 13-15), 143 (fraction 28–30), 231 (fraction 31-33), 223 (fraction 40-42), 212 (fraction 43–45) and 66 (fraction 52-54) candidate taste-active peptides in the fractions, respectively, comprising 266 peptide sequences.

[0073] MaxQuant software (Max Planck Institute of Biochemistry in Martinsried, Germany) was used to determine peptide sequences using the methodology of Jürgen Cox et al., " MaxQuant enables high peptide identification rates, individualized p.p.b.-range mass accuracies and proteome-wide protein quantification," Nat. Biotechnol., 26(12); 1367–72 (2008). In certain aspects, the MaxQuant software creates a list of all possible peptides that could theoretically be formed from the specified proteins. In this case, bovine alpha si-, alpha s2-, beta- and kappa-casein were specified, since they are the most abundant proteins in cheese. The protein sequences can be found in the database UniProt. MaxQuant simulates the peptide fragmentation pattern, which is highly predictable, and compares these obtained theoretical mass spectra with the measured LC-ToF-MS data. The output is a list of peptides that were found in the measured LC-ToF-MS mass spectra, and the score states how close the match was between the theoretical and the measured spectra. Therefore, a high score makes it more likely that the peptide is contained in the original sample. The cut-off value that was used to predict whether a peptide is contained in a sample was a score of 50, and, if the score is over 100, the peptide is assumed to be almost certainly contained in the sample.

[0074] The source protein sequences were obtained from the UniProt database. The most abundant milk proteins— bovine αs1- (ID P02662), αs2- (ID P02663), β- (ID P02666), and κ- (ID P02668) casein— were imported into the MaxQuant software for in silico unspecific digestion and comparison with the LC-ToF-MS data. MaxQuant analysis was run with LC-ToF-MS data, once for an intact sample of water-soluble extract (from classic extraction) in water, and also for aliquots of each MPLC fraction obtained after fractionation of the water-soluble extract.

[0075] The analysis of the cheddar cheese via MaxQuant software revealed the presence of 819 candidate taste-active peptides, with 438 of these peptides having a score of 50 or higher, and 126 of these peptides having a score of 100 or higher. In this case, the MaxQuant software found 819 peptide sequences in all MPLC fractions with the specified parameters (chain length of 2 to 25 amino acids, origin protein sequences of bovine αs1-, αs2-, β- and κ-casein, unspecific digestion, contaminants not included). The phrase "candidate taste-active peptides" in reference to the 819 peptides revealed by the Max Quant software means that these 819 peptides could be found in sufficient amounts for detection based on the search parameters and may be taste¬ active. In some aspects, the connection of the peptides to "taste" may be obtained by analyzing the peptides within the fractions with the highest (comparative) taste dilution factor, which are expected to include the most relevant taste-active peptides.

[0076] Using targeted LC-MS / MS analysis, the identity of peptides postulated by the MaxQuant software was confirmed based on the alignment of multiple reaction monitoring (MRM) transitions characteristic for each peptide. The amino acid sequences of the 266 candidate peptides were entered into Skyline Software (a free and open-source tool), followed by MRM computation. This software uses the predictable fragmentation patterns of the peptides, which are characteristic for each input peptide sequence, to create LC-MS / MS methods for targeted peptide analysis (MacLean et al., 2010). The MRM transitions were exported to multiple T.. C-MS / MS methods with a maximum of 150 MRM transitions per method and 5 ms d’well time at unit resolution for QI and Q3. If a peptide exists in a sample, the MRM transitions align in one chromatographic signal at the same retention time. To ensure reliable peptide identification, peptides were only considered as confirmed when at least five MRM transitions aligned in one chromatographic signal[00771 Then LC-MS / MS analysis was performed using the mass transitions computed by Skyline and a standardized chromatography procedure. The LC-MS / MS system comprised a Shimadzu Nexera X2 ultra-high-performance liquid chromatography (UHPLC) system (Shimadzu, Kyoto, Japan), connected to a 5500 QTrap mass spectrometer (AB Sciex, Darmstadt, Germany). Targeted analysis was then performed via liquid chromatography coupled with tandem mass spectrometry (LC-MS / MS) following the method adapted from Sebald et al., 2018. Samples were cooled in an autosampler at 10 °C and 5 pL aliquots were injected into the LC-MS / MS system for analysis. The UHPLC was equipped with a Kinetex C 8 column (1.7 pm, 100 A, 100 x 2.1 mm, Phenomenex, Aschaffenburg, Germany) heated at 40 °C in a column oven, and was operated using 0.1 % (v / v) formic acid in water (A) and 0.1 % (v / v) formic acid in acetonitrile (B) as solvents. At a flow rate of 0.4 mL / min, the separation gradient was as follows: 5% B held constant for 1 min, increased to 100% B within 11 min, held constant for 3 min, decreased to 5% B within 1 min and finally held constant for 2 min. The analytes were ionized in positive electrospray ionization (ESI) and detected in MRM mode. The ionization voltage was 5500 V and the source temperature 450 °C with nitrogen as curtain gas at 35 psi and heater gas at 65 psi, and air as nebulizer gas at 55 psi.

[0078] To prepare the samples for targeted peptide analysis, a portion of the crude cheese extract (0.3 g) was re-dissolved in water (1 mL), as described above. The mixture was vortexed and shaken for 1 hour at room temperature, then centrifuged and membrane-filtered (0.45 pm) to remove any insoluble residues prior to injection into the LC-MS / MS system.

[0079] The mass spectrometer can distinguish between the different peptides. Targeted LC-MS / MS analysis of the crude extract from cheddar cheese revealed that for 204 of the 266 candidate taste-active peptides, the MRM transitions aligned in one chromatographic signal. Therefore, it was concluded that these peptides were contained in the cheddar cheese crude extract. The remaining peptides showed no alignment of the MRM transitions and were excluded in further steps.

[0080] The list of candidate peptides was further reduced to 184 peptides by identifying the peptides having a signal to noise ratio of > 10 in the recorded targeted LC-MS / MS data. The signal to noise ratios (S / N) were calculated from the targeted LC-MS / MS data as the peak area of the peptide divided by the area of the noise. The peak areas can be determined for each MRM transition, so characteristic mass transitions were selected for each peptide to do this. Generally, the signal- to -noise ratio facilitates distinguishing meaningful data (signal) from backgro und interference or irrelevant data (noise) and helps identify and quantify taste-active peptides in a complex mixture, such as cheese. This filter ensured a sufficiently high abundance of the peptides in the cheddar cheese for a contribution to the cheese taste.100811 Fold-Change Analysis2.5Attorney Docket 1410-163200-PC

[0082] Next, the fold-change was calculated as an indicator of differences between two samples. Fold-change refers to the relative difference in the concentration (or amount) of a specific peptide between two samples with different sensory properties. Here, fold-change was determined between two cheddar cheese samples (named for purposes of this experiment " Sample A" and " Sample B") with different salty taste intensities and ripening times. Sample A was the cheddar cheese used in the extraction and sensory analysis described above. Sample A had a 12-month ripening time, and Sample B was a cheddar cheese with a 3-month ripening time. Selection of these samples was based on the assumption that longer ripening times coincide with a higher degree of proteolysis and, subsequently, increased amounts of peptides that impact the salty taste of the cheese. Sample B was also perceived as less salty than Sample A during taste analysis.

[0083] With respect to fold-change, the targeted LC-MS / MS data was used. The foldchange metric is used to assess how much the concentration of a particular peptide changes between different conditions, samples, or time points, and is often used to identify peptides that are most responsible for the flavor profile, in LC-MS analysis, when a sample is analyzed, different peptides will elute and ionize at different times, generating distinct peaks in a chromatogram, and the area under the peak is proportional to the amount or concentration of that peptide in the sample. For calculation of the fold -change, the peak area ratio is the ratio of the peak area of a target peptide in two samples with different properties. The ratio gives an indication of how much more (or less) of a particular peptide is present in one sample relative to another.

[0084] In the context of taste-active peptides m cheese, fold-change helps to identify which peptides increase or decrease in concentration between different cheese samples, such as cheeses that are aged differently, processed differently, or prepared under different conditions. Peptides with a high fold-change (i.e., significantly increased peak area in one sample compared to another) could be identified as contributing more to the flavor profile of the cheese and may explain any differences in the sensory profiles of the two cheese samples. These are potentially the most taste-active peptides that play a major role in flavor perception. Peptides with a fold¬ change around 1 might not significantly affect the flavor differences between the samples and may be considered less important in the context of flavor development. Furthermore, a fold-change of less than 1 suggests that the peptide of interest is more abundant in the cheese sample with sensory properties that differ from the desired taste profile.

[0085] For example, if a peptide's peak area in one sample (e.g., cheese from a certain agin? process! is twice as large as in another sample fe.g.. cheese from a different aging process), the fold-change would be 2 (or a 2-fold increase). Conversely, if the peak area in one sample is half of the other, the fold-change would be 0.5 (or a 2-fold decrease).

[0086] Fold-change is calculated as follows:Fold-change = (peak area in condition A) / (peak area in condition B).

[0087] Using the fold-change peak area ratio, the number of candidate peptides was further reduced to 25 peptides by picking out the peptides with a fold-change of 4 or above. The chosen fold-change of 4 suggests that there is a relevant difference in peptide concentrations between the two samples. Since the peptides of interest were the peptides that cause the increased salty taste in Sample A cheese compared to Sample B cheese, the focus was on the peptides that had a large difference in concentrations. Setting the cutoff fold-change value at 4 ensured a high likelihood that the observed sensory properties were not simply caused by natural fluctuations.

[0088] After reducing the number of taste-modulating candidates to 25, the 25 peptides were custom synthesized by Genscript (New Jersey, USA). Based on quantitative ¹H-NMR spectroscopy, the resulting peptides were 71-96% pure. Furthermore, targeted LC-MS / MS analysis using the same method described above confirmed the identity of the 25 candidate peptides by close agreement of retention times and the signal intensity ratios of the MRM transitions. Therefore, it could be confirmed that all 25 candidate peptides occurred in the investigated Cheddar cheese (Sample A).[00891 The reference compounds of the 25 candidate peptides were then separately evaluated by a trained sensory panel to assess the taste properties of each peptide to determine which ones have significant taste activity, such as sweetness, bitterness, umami, etc. In particular, the intrinsic taste thresholds of the candidate peptides in water were assessed to evaluate astringent, bitter, and sour taste peptides.27Attorney Docket 1410-163200-PC

[0090] Samples were prepared of each of the 25 peptides in water at a beginning concentration of 1 mmol / L and diluted stepwise 1:2 with water to a total of up to 24 dilution steps until the panelists could no longer perceive a difference between the peptide solution and water. Before sensory evaluation, the pH value was adjusted to 5.4 using trace amounts of formic acid or sodium hy droxide to represent the pH of the cheese. The samples were evaluated by a trained sensory panel comprised of twelve trained panelists. Each panelist tasted the samples in a set of duo-trio tests in increasing concentrations, identified the sample that deviated from the two blanks (water), and stated which taste quality was detected. The procedure followed here was the same as the procedure described above for TDA and cTDA except that single compounds at known concentrations were used here instead of analyte mixtures with unknown compositions. Similarly to the methodology used for TDA and cTDA, the panel taste threshold was calculated as the geometric mean of the individual taste thresholds for each panelist.

[0091] Further evaluation of the 25 peptides was performed in the model broth system. The peptides were evaluated by the sensory panel for modulation of any one or more taste qualities they perceived in a sample. Samples were prepared of each of the 25 peptides in model broth at a beginning concentration of 1 mmol / L and diluted stepwise 1:2 with model broth to a total of up to 30 dilution steps until the panelists could no longer perceive a difference between the peptide solution and model broth. Before sensory evaluation, the pH value was adjusted to 5.4 using trace amounts of formic acid or sodium hydroxide to match the pH of the starting cheese (Sample A). Again, the samples were evaluated by a trained sensory panel comprised of twelve trained panelists. Each panelist tasted the samples in a set of duo-trio tests in increasing concentrations, identified the sample that deviated from the two blanks (model broth) and stated which taste quality was detected. The panel taste threshold was calculated as the geometric mean of the individual taste thresholds for each panelist.

[0092] The intrinsic taste thresholds (bitter, astringent, sour) in water of the 25 candidate compounds are provided in Table 4 below. In this experiment, the taste threshold value means that a bitter, astringent, or sour taste was first identified by the tasting panel at the concentration listed in Table 4. The taste- odulating thresholds (salty, bitter umami, kokumi, astringent) in the model broth system are also provided in Table 4. Peptides that have a higher 28Attorney Docket 1410-163200-PCtaste threshold value for bitter, astringent, or sour taste are generally more advantageous for use as a taste modifier because they can be used in higher amounts to contribute a desired taste (e.g., sally or umami) but without adding an undesirable taste (e.g., bitter, astringent, or sour) at that concentration.

[0093] Table 4 - Intrinsic Taste Threshold and Taste Modulating Threshold of PeptidesPeptides Intrinsic Taste Taste Amount in Threshold in Threshold in T hreshold in Starting (amino acidWater Model Broth Model Broth Cheddar sequences)< Intrinsic Cheese Threshold inWaterE1V 20.3 mg / L 7.4 mg / L Yes 0.05 mg / kg (bitter) (salty)(56.6 μmol / L) (20.6 μmol / L)YGLN (SEQ ID NO. 40.6 mg / L. 47.4 mg / L No 0.94 mg / kg 2) (bitter) (sally)(87.2 gmoi / L) (101.9iimol / L)NIPPLTQTPV (SEQ 34.9 mg / L. 32.7 mg / L 4.65 mg / kg ID NO. 3) (bitter) (bitter)(32.4 μmol / L) (30.3 μmol / L)AVFYPQ (SEQ ID 38.6 mg / L 73.4 mg / L No 13.14 mg / kg NO. 4) (astringent) (salty)(57.2 μmol / L) (109 μmol / L)RDMPIQA (SEQ ID 119.7 mg / L 49.4 mg / L Yes 34.54 mg / kg NO. 5) (sour) (umami)(144.2 μmol / L) (59.6 μmol / L)imioi / L)QEPVLGPVRGPFP 179.1 mg / L 24.4 mg / L Yes 16.23 mg / kg (SEQ ID NO. 6) (bitter) (umami)(128.6 μmol / L) (17.5 μmol / L)imioi / L)NIPPLTQ (SEQ ID 37.1 mg / L 48.7 mg / L No 1.84 mg / kg NO. 7) (bitter) (kokumi)(47.5 μmol / L) (62.2 μmol / L)29Attorney Docket 1410-163200-PCPeptides intrinsic Taste Taste Amount in Threshold in Threshold in Threshold in Starting (amino addWater Model Broth Model Broth Cheddar sequences)< Intrinsic Cheese Threshold inWaterHQFHQPLFP (SEQ 125.8 mg / L 18.2 mg / L Yes 7.02 mg / kg ID NO. 8) (sour) (salty)(119.8 μmol / L) (17.3 μmol / L)μmol / L)PGPI (SEQ ID NO. 9) 26.0 mg / L 7.0 mg / L Yes 0.16 mg / kg (bitter) (umami)(68.1 μmol / L) (18.2 μmol / L)RDMPIQ (SEQ ID 32.0 mg / L 31.9 mg / L Slightly 5.10 mg / kg NO. 10) (bitter) (salty)(42.2 μmol / L) (42 μmol / L)NAVPITFT (SEQ ID 86.6 mg / L 165.2 mg / L No 4.16 mg / kg NO. 11) / bitter) (salty)(106.7 (203.4pmol / L) umol / L)SKVLPVPQ (SEQ ID 82.7 mg / L 41.3 mg / L Yes 6.30 mg / kg NO. 12) (bitter) (95.4 (salty)pmol / L)(47.7 pmol / L)GPVRGPFPIIV (SEQ 37.2 mg / L 8.7 mg / L * 7,96 mg / kg ID NO. 13) (bitter) (bitter)(32.3 μmol / L) (7.6 μmol / L)EEIVPN (SEQ ID 46.5 mg / L. 22.8 mg / L * 2.13 mg / kg NO. 14) (sour) (bitter)(66.4 μmol / L) (32.6 μmol / L)VVPP (SEQ ID NO. 36.3 mg / L 38.9 mg / L No 1.81 mg / kg 15) (bitter) (salty)(88.3 μmol / L) (94.8 pmol / L)AVRSPAQIL (SEQ 17.2 mg / L 78.2 mg / L No 0.69 mg / kg ID NO. 16) (bitter) (sally)(18 μmol / L) (82 μmol / L)FVAP (SEQ ID NO. 1.3 mg / L 29.5 mg / L No 7.32 mg / kg17) (bitter) (salty)30Attorney Docket 1410-163200-PCPeptides Intrinsic Taste 'Paste Amount in Threshold in Threshold in Threshold in Starting (amino addWater Model Broth Model Broth Cheddar sequences)< Intrinsic Cheese Threshold inWater(29 pmol / L) (68.1 jimol / L)SLPQNIPP (SEQ ID 2.2 mg / L 49.7 mg / L No 2.36 mg / kg NO. 18) (astringent) (salty)(2.5 pmol / L) (57.4 jimol / L)VLGPVR (SEQ ID 11.6 mg / 1. 9.6 mg / L Slightly 0.39 mg / kg NO. 19) (umami)(astringent)(15 pmol / L)(18.1 pmol / L)NIPPL (SEQ ID NO. 39.7 mg / L 57.1 mg / L No 2.46 mg / kg 20) (bitter) (bitter)(71.9 pmol / L) (103.4gmol / L)KAVPYPQ (SEQ ID 81.6 mg / L 54.5 mg / L Yes 16.33 mg / kg NO. 21) (bitter) (umami)(101.7 (68 gmol / L)pmol / L)QEPVL (SEQ ID NO. 70.7 mg / L 81.9 mg / L No 2.56 mg / kg 22) (bitter) (astringent)(120.9 (140.1pmol / L) gmol / L)DMPIQA (SEQ ID 43.1 mg / L 81.3 mg / L No 7.70 mg / kg NO. 23) (bitter) (umami)(64 iimol / L) (120.7gmol / L)KVLPVPQ (SEQ ID 13.0 mg / L 47.4 mg / L No 19.09 mg / kg NO. 24) (bitter) (salty)(16.7 imioi / L) (60.8 gmol / L)VLGPVRGPFP (SEQ 20.3 mg / L 86.6 mg / L No 0.73 mg / kg ID NO. 25) (bitter) (umami)(19.6 umol / L) (83.4 pmol / L)* The taste threshold in model broth was lower than the intrinsic threshold m water but the taste was a bitter taste.31Attorney Docket 1410-163200-PC

[0094] The intrinsic taste thresholds ranged between 2.5 μmol / L to 144.2 μmol / L in water. The taste-modulating thresholds ranged between 7.6 and 203.4 μmol / L in the model broth. In this experiment, the taste threshold value means that a salty, bitter, umami, kokumi, sour, or astringent taste was first identified at the concentration listed in Table 3. The peptides having; a taste modulating threshold concentration in model broth for salty, umami, or kokumi that was lower than an intrinsic taste threshold concentration in water for bitter, astringent, or sour were deemed the most promising taste modulating peptides.100951 The taste threshold values in model broth for each peptide were also compared to the amount of each peptide in the starting cheddar cheese. The amounts of the peptides in the cheddar cheese were generally much lower than the taste threshold values in the model broth. Despite the peptides being present in the starting cheddar cheese in amounts lower than would appear to be necessary to taste them on an individual basis, the results of Example 2 indicate that the salty-modulating peptides enhance saltiness in certain food or beverage systems at amounts lower than the taste threshold in model broth.{0096] In Table 4, there are twelve peptides that were determined to have salty-modulating activity, namely: EIV, YGLN (SEQ ID NO. 2), AVPYPQ (SEQ ID NO. 4), HQPHQPLPP (SEQ ID NO. 8), RDMPIQ (SEQ ID NO. 10), N AV PITPT (SEQ ID NO. 11), SKVLPVPQ (SEQ ID NO. 12), VVPP (SEQ ID NO. 15), AVRSPAQIL (SEQ ID NO. 16), FVAP (SEQ ID NO. 17), SLPQNIPP (SEQ ID NO. 18), and KVLPVPQ (SEQ ID NO. 24). The salty- modulating thresholds of each of these twelve candidate peptides were compared to their respective intrinsic taste thresholds (i.e., bitter, astringent, and sour) to identify which of these twelve peptides have a taste modulating threshold concentration for salty that is lower than their respective intrinsic taste threshold.

[0097] As a result, it was determined that the following three out of twelve salty- modifying peptides have salty-modulating thresholds that are lower than their respective intrinsic taste threshold: EIV (taste modulating threshold (salty) 20.6 μmol / L and intrinsic threshold (bitter) 56.6 μmol / L): HQPHQPLPP (SEQ ID NO. 8) (taste modulating threshold (salty) 17.3 μmol / L and intrinsic threshold (sour) 119.8 μmol / L); and SKVLPVPQ (SEQ ID NO.12) (taste modulating threshold (salty) 47.7 μmol / L and intrinsic threshold (bitter) 95.4 μmol / L).

[0098] This means that each of these three candidate peptides exhibits a salty-modifying taste when it is present in a sample at a concentration that is lower than the concentration of the peptide that is required to exhibit the intrinsic (bitter, sour, or astringent) taste of this peptide in a sample. Such peptides are desirable for use to modulate the salty taste of a food or beverage product. To enhance the salty taste of a given food or beverage product, a flavor enhancing composition including one or more of the three salty-modifying peptides EIV, HQPHQPLPP (SEQ ID NO. 8), and SKVLPVPQ (SEQ ID NO. 12) may be added to the food or beverage product to impart a salty taste without negatively affecting the overall flavor of the food or beverage product (e.g., by adding bitter, sour, or astringent flavor notes to the food or beverage product).

[0099] On the other hand, the other nine candidate peptides had a taste modulating threshold concentration for salty that is higher than or very close to its respective intrinsic taste threshold (bitter, sour, astringent): YGLN (SEQ ID NO. 2) (taste modulating threshold (salty) 101.9 μmol / L and intrinsic threshold (bitter) 87.2 μmol / L); AVPYPQ (SEQ ID NO. 4) (taste modulating threshold (salty) 109 μmol / L and intrinsic threshold (astringent) 57.2 μmol / L); RDMPIQ (SEQ ID NO. 10) (taste modulating threshold (salty) 42 μmol / L and intrinsic threshold (bitter) 42.2 μmol / L); NAVPITPT (SEQ ID NO. 11) (taste modulating threshold (salty) 203.4 μmol / L and intrinsic threshold (bitter) 106.7 μmol / L); VVPP (SEQ ID NO. 15) (taste modulating threshold (saltv) 94.8 μmol / L and intrinsic threshold (bitter) 88.3 μmol / L);AVRSPAQIL (SEQ ID NO. 16) (taste modulating threshold (salty) 82 μmol / L and intrinsic threshold (bitter) 18 μmol / L); FVAP (SEQ ID NO. 17) (taste modulating threshold (salty) 68.1 pmol / L and intrinsic threshold (bitter) 29 pmol / L); SLPQNIPP (SEQ ID NO. 18) (taste modulating threshold (salty) 57.4 μmol / L and intrinsic threshold (astringent) 2.5 μmol / L; and KVLPVPQ (SEQ ID NO. 24) (taste modulating threshold (salty) 60.8 μmol / L and intrinsic threshold (bitter) 16.7 μmol / L). This means that each of these other nine salty-modifying peptides exhibit a bitter, sour, or astringent taste in a sample at a concentration that is lower than the concentration required of this peptide to modify the salty taste of a sample. At least in some approaches, such peptides may not be desirable for use to modulate the salty taste of a 33Attorney Docket 1410-163200-PCfood or beverage product. It will be appreciated, however, that such peptides may be added to a food and may interact with other substances / food matrix to favorably change the taste perception of the food product.

[0100] Example 2100101 ] Evaluation of Salty-Enhancing Peptides in Model Food Systems

[0102] Model systems for the following food product types were evaluated: process cheese (sauce form), cream cheese (plain), plant-based cream cheese (plain), sauces (soy sauce and tomato frito), condiments (tomato ketchup), gravy (ready-to-use), and soup (chicken broth). Each of the three salty -modifying peptides EIV, HQPHQPLPP (SEQ ID NO. 8), and SKVLPVPQ (SEQ ID NO. 12) was separately added to each system in different concentrations and evaluated for salty intensity by a trained human sensory panel.

[0103] Prior to evaluating samples containing salt-modifying peptides, a human sensory panel was first trained on sodium chloride (NaCl) solutions as a control. Reference lines of NaCl concentrations (30 mM, 35 mM, 40 mM, 45 mM, 50 mM, 55 mM, 60 mM, 65 mM, 70 mM, and 75 mM) were prepared in water and model broth. The model broth (1.9 g monosodium L- glutamate, 6.4 g maltodextrin, and 2.1 g yeast extract in 1 L water) was initially prepared without sodium chloride and spiked with NaCl to obtain the respective NaCl concentration. In multiple sessions, coded samples of NaCl concentrations (samples A-D) were prepared in water and model broth, respectively. Panelists were asked to taste the samples and match each sample to the NaCl reference line in water or model broth, respectively. Those sessions were conducted until each reference concentration was tested as a coded sample at least twice. This methodology was used to train panelists on iso-intensity evaluations and to identify the confidence interval of each NaCl concentration in water and model broth.

[0104] Another set of NaCl concentrations (5 mM, 10 mM, 15 mM, 20 mM, 30 mM, 50 mM, 65 mM, 80 mM, 100 mM, and 150 mM) was prepared in the model broth excluding prior NaCl addition. These samples were prepared as coded samples A-J in order of increasing concentration. Panelists were asked to evaluate the perceived salty intensity on a scale without limitation and without predetermined values ("magnitude estimation" approach). Theindividual dose-response curves were normalized to 50 mM (which corresponds to the "evaluated salty taste intensity" of 5) preserving the individual rating ratios of each panelist. This methodology was used to identify a dose-response curve tor NaCl in the model broth tor comparison with further recorded dose-response curves.

[0105] Next, to determine their salty enhancing potential in the model broth system, stock solutions were prepared of each of the three salty -modifying peptides (EIV, HQPHQPLPP (SEQ ID NO. 8), and SKVLPVPQ (SEQ ID NO. 12)). The aim of the present experiment was to identify the lowest concentration of peptide at which there is the greatest salty enhancing effect. A model broth was prepared by mixing 29 g NaCl (~ 50 mM), 1.9 g monosodium L-glutamate, 6.4 g maltodextrin, and 2.1 g yeast extract in water (1 L), and the pH value was adjusted to 5.4 using trace amounts of formic acid or sodium hydroxide (to provide the approximate pH of cheddar cheese). Each of the three salty-modifying peptides were dissolved in the model broth at a concentration four times (4x) the respective taste threshold of each salty-modifying peptide in model broth shown in Table 4. The stock solutions were then diluted with model broth: 1:2, 1:2.66, 1:4, and 1:8 (i.e., 2x, 1.5x, lx, and 0.5x the taste threshold in model broth, respectively).

[0106] The trained sensory panel comprised of twelve trained panelists was presented with a reference sample (prepared model broth) and five coded samples (samples A-E). The salty taste intensity of the reference sample was designated as a "5" to correlate with a concentration of 50 mM NaCl in the model broth. The coded samples were evaluated by the panel. The panelists were asked to taste each sample and assign it a value regarding its saltiness in direct correlation to the reference sample. The panelists were instructed that a value greater than 5 is representative of a more intense salty taste than the reference, while a value less than 5 is representative of a less intense salty taste than the reference. The salty taste intensity values of the three candidate taste-modifying peptides in model broth identified by the panelists are provided in Tables 5A-5C below. These values are an average of 9-12 separate values determined by the panelists.

[0107] The theoretical ability of each peptide to be used for sodium reduction can be calculated based on the observed salty taste enhancement The model broth without a spiked peptide has a " Baseline Salty Taste Intensity" of 5.00. Therefore, a " Salty Enhancement" and" Relative Sodium Reduction" percentage values can be calculated according to the following formulas:" Salty Enhancement %" = (Evaluated Salty Taste Intensity ■■ Baseline Salty Taste Intensity) / Baseline Salty Taste Intensity‘'' Relative Sodium Reduction %" = (Evaluated Salty ’Taste Intensity - Baseline Salty Taste Intensity) / Evaluated Salty Taste Intensity

[0108] Because the model broth (unspiked) has a Baseline Salty Taste Intensity value of 5.00, when a peptide provides an Evaluated Sally Taste Intensity value of greater than 5.00, the amount of sodium in a food or beverage system containing the peptide theoretically could be reduced to the amount needed to provide a value of 5.00 without any loss of salty perception.

[0109] The average of 9–12 values provided by the panelists for salty taste intensity are listed in Tables 5A-5C. The calculated Salty Enhancement Percentage and Relative Sodium Reduction Percentage for the three salty taste-modifying peptides in model broth are also provided in Tables 5A-5C below.

[0110] Table 5A – Salty Taste Intensity of EIV Peptide in pH-Adjusted Model Broth Peptide Cone, EIV Peptide(x threshold in Cone. Evaluated Salty Relativemodel broth) (μmol / L) Salty Taste- Enhancement SodiumIntensity Reduction0.5x 10.3 6.15 n / a - 1x 20.6 5.05 n / a1.5x 30.9 6.75 35% 26%2x 41.2 8.85 77% 44%4x 82.4 8.55 71% 42%

[0111] As determined in Example 1, the EIV peptide had a modulating taste threshold (salty) in model broth of 20.6 μmol / L and an intrinsic taste threshold in water of 56.6 μmol / L (bitter taste). As shown in Table 5A, the EIV peptide clearly enhanced the salty perception with increasing levels in pH-adjusted model broth. The strongest salty enhancing effect was found at 2x threshold concentration and the impact of the EIV peptide on salty perception may level out at around 2x threshold concentration, as the salty enhancing effect was slightly lower at 4x36Attorney Docket 1410-163200-PCthreshold concentration. It was found that spiking the model broth with 2x the modulating taste threshold may enable up to 44 % sodium reduction.

[0112] Table 5B - Salty Taste Intensity of HQPHQPLPP Peptide (SEQ ID NO. 8) in Model BrothPeptide Cone, HQPHQPLPP Peptide (SEQ ID NO. 8)(x threshold in Cone. Evaluated Salty Relativemodel broth) (μmol / L) Salty Taste Enhancement SodiumIntensity Reduction0.5x * *lx 17.3 6.37 27% 22%1.5x 25.95 6.99 40% 28%2x 34.6 7.81 56% 36%4x 69.2 7.69 54% 35%

[0113] As determined in Example 1, the HQPHQPLPP peptide (SEQ ID NO. 8) had a modulating taste threshold (salty) in model broth of 17.3 μmol / L and an intrinsic taste threshold in water of 119.8 μmol / L (sour taste). As shown in Table 5B, the HQPHQPLPP peptide (SEQ ID NO. 8) clearly enhanced the salty perception w ith increasing levels in pH- adjusted model broth. The strongest salty enhancing effect was found at 2x threshold concentration and the impact of the HQPHQPLPP peptide (SEQ ID NO. 8) on salty perception may level out at around 2x threshold concentration, as the salty enhancing effect was no greater at 4x threshold concentration than at 2x threshold concentration. It w as found that spiking the model broth with 2x the modulating taste threshold may enable up to 36% sodium reduction.

[0114] Table 5C - Salty Taste Intensity of SKVLPVPQ Peptide (SEQ ID NO. 12) in Model BrothPeptide Cone, SKVLPVPQ Peptide (SEQ ID NO. 12)(x threshold in Cone. Evaluated Salty Relative Sod iummodel broth) (pmol / L) Salty Taste Enhancement ReductionIntensity0.5x * * *lx 47.7 7.23 45% 31%1.5x 71.6 8.13 63% 38%95.4 8.25 65% 39%4x 190.8 9.25 85% 46%Attorney Docket 1410-163200-PC

[0115] As determined in Example 1, the SKVLPVPQ peptide (SEQ ID NO.12) had a modulating taste threshold (salty) in model broth of 47.7 umol / L and an intrinsic taste threshold in water of 95.4 pmol / L (bitter). As shown in Table 5C, the SKVLPVPQ peptide (SEQ ID NO. 12) clearly enhanced the salty perception with increasing levels in pH-adjusted model broth. Of all three peptides, the SKVLPVPQ peptide (SEQ ID NO. 12) had the highest salty impact at low dosage (lx). The strongest salty enhancing effect was found at 4x threshold concentration, and it was found that spiking the model broth with 4x the modulating taste threshold may enable up to 46% sodium reduction. It is possible that salty intensity may be increased beyond the 4x threshold concentration.

[0116] Model Food Systems

[0117] The salty-modifying peptides were then evaluated in model food systems (cheese sauce, cream cheese, soy sauce, ketchup, plant-based cream cheese spread, tomato frito, chicken broth, and turkey gravy). The cheese sauce included about 16.8% fat, 62.3% moisture, and 2.3% salt, and had a pH of about 5.7. The cream cheese included about 26% fat 63.3% moisture, and 0.8% salt, and had a pH of about 4.95. The soy sauce included about 83% moisture, about 0.8% salt, and had a pH of about 5.1. The tomato ketchup included about 68.7% moisture and 2.2% salt, and had a pH of about 3.7. The plant-based cream cheese included 15.5% fat, 77.0% moisture, about 1.2% salt, and had a pH of about 4.2. ’The gravy included about 1.5% fat, 93.7% moisture, and 0.9% salt, and had a pH of about 5.5. The tomato frito included about 2.0% fat, 89.0% moisture, and 7.2% salt, and had a pH of about 4.25. The chicken broth included about 97.7% moisture, and 0.61% salt, and had a pH of about 7.34.

[0118] Salt-reduced versions of each food product were also prepared. The salt contents of these formulations were reduced by about 20%.

[0119] To evaluate the model systems, the model food systems were prepared by adding the three taste-mod ifying peptides (EIV (SEQ ID NO. 1), HQPHQPLPP (SEQ ID NO. 8), or SKVLPVPQ (SEQ ID NO. 12)) into the salt-reduced formulations two to three days before the evaluation. The peptides were added in concentrations of two times (2x) and four times (4x) their respective taste thresholds in model broth (as shown in Table 4). Panelists were thoroughly trained on iso-intensity evaluations in a concentration range of 30 mM to 75 mM,38Attorney Docket 1410-163200-PCwhich was also the range in which the panelists were most able to detect saltiness while also not being overwhelmed by the saltiness. Before sensory evaluation, the salt-reduced formulations were diluted with milk or natural spring water to obtain a perceived saltiness in the trained concentration range of 30 mM to 75 mM NaCl in water. 'This enabled the panelists to consistently evaluate the salt-reduced formulation and to evaluate a possible increase in saltiness of the spiked formulations.

[0120] A team of eight panelists were presented with a reference line of 30 mM to 75 mM NaCl in water (in 5 mM increments), a control sample (equivalent to one of the concentrations in the reference line), and four coded samples (samples A-D): (A) original formulation; (B) salt-reduced formulation; (C) salt-reduced formulation with evaluated taste¬ modifying peptide at a concentration of two times (2x spiked) the taste threshold in model broth; and (D) salt-reduced formulation with evaluated taste-modifying peptide at a concentration of four times (4x spiked) the taste threshold in model broth.

[0121] The panelists were asked to taste each sample and pinpoint an isointense saltiness with respect to the presented reference line. Panelists were not limited by the range of the reference line and were allowed to evaluate foe samples outside of this range. The evaluated salty intensities are represented as an average. The salty taste intensity of the three candidate taste-modifying peptides in model systems are provided in Tables 6-8 below.

[0122] Tables 6, 7, and 8 include the results of the EIV peptide, the HQPHQPLPP peptide (SEQ ID NO. 8), and the SKVLPVPQ peptide (SEQ ID NO. 12) in the model food systems. As used herein, " TC" refers to the threshold concentration for "salty" taste, "2x” refers to a concentration that is two times the threshold concentration, and "4x" refers to a concentration that is four times the threshold concentration.

[0123] Table 6 -- E1V Peptide in Model Food SystemsTC / Diluted Salty Taste Intensity2x TC / TC / 2x Control - Sample C Sample D Control ~Model 4 x TC / 4x Sample (2x (4x Dilution Sample ASystem (μmol / L) (μmol / L) B (Salt Spiked Spiked (Originalin Model Reduced) w / with Formulation)Broth Peptide) Peptide) 20.6 n / a41.2 5.9Cheese 1:6 in(0.29x TC) 61.82 49.60 57.36 62.70 sauce milk82.4 11.8(0.57x TC)20.6 n / aCreamNone 41.2 n / a 63.70 53.71 64.50 68.80 cheese82.4 n / a20.6 n / a41.2 1.37Soy 1:29 in(0.07x TC) 53.05 44.15 48.33 40.23 sauce water82.4 2.8(0.13x TC)20.6 n / a41.2 5.9Tomato 1:6 in(0.29x TC) 43.25 39.80 52.44 50.19 ketchup water82.4 11.8(0.57x TC)20.6 n / aPlant41.2based 1:3 in 10.363.83 52.60 58.95 55.7 cream water (0.5 x TC) 3 cheese 82.4 20.6(lx TC)20.6 n / a41.2 10.3Tomato 1:3 in(0.5x TC) 53.00 40.63 53.18 47.60 frito water82.4 20.6(lx TC)20.6 n / a41.2 20.6Turkey 1:1 in(Ix TC) 67.63 56.32 57.22 59.25 Gravy water82.4 41.2(2x TC)20.6 n / a1:1 in 41.2 20.6Chickenchicken (lx TC) 41.41 38.66 42.81 40.30 brothbroth 82.4 41.2(2x TC)40Attorney Docket 1410-163200-PC

[0124] Table 7 - HQPHQPLPP Peptide (SEQ ID NO. 8) in Model Food SystemsTC / Diluted Salty Taste Intensity2x TC / TC / 2x / 4x Control - Sample C Sample D Control - Model 4x TC (μmol / L) Sample (2x (4x Spiked Dilution -) Sample ASystem (μmol / L) B (Salt Spiked w / in Model (OriginalReduced) w / Peptide) Broth Formulation)Peptide)17.3 n / a34.6 4.9Cheese1:6 milk (0.29x TC) 61.20 5295 56.58 61.19 sauce69.2 9.9(0.57x TC)17.3 n / aCreamNone 34.6 n / a 57.57 45.92 60.30 61.95 cheese69.2 n / a17.3 n / a34.6 1.2Soy 1:29(0.07x TC) 54.45 47.43 50.83 50.88 sauce water69.2 2.3(0.13x TC)17.3 n / a34.6 4.9Tomato 1:6(0.29x TC) 40.69 47.49 48.54 51.75 ketchup water69.2 9.9(0.57x ’FC)17.3 n / aPlantbased 34.6 8.71:3(0.5x TC) 58.60 52.53 55.15 55.97 cream watercheese 69.2 17.3(17.3x TC)17.3 n / a34.6 8.7Tomato 1:3(0.5x ’FC) 54.58 46.55 53.53 50.08 frito water69.2 17.3(lx TC)17.3 n / a34.6 17.3Turkey 1:1(lx TC) 65.08 57.55 60.38 59.68 Gravy water69.2 34.6(2x TC)17.3 n / a1:1 in 34.6 17.3Chickenchicken (lx TC) 46.79 41.74 49.29 45.50 brothbroth 69.2 34.6(2x TC)41Attorney Docket 1410-163200-PC

[0125] Table 8 - SKVLPVPQ Peptide (SEQ ID NO. 12) in Model Food SystemsTC / Diluted TC Salty Taste Intensity2x TC / / 2x / 4x Control - Sample C Sample D Control - Model 4x TC (μmol / L)Dilution Sample (2x Spiked (4x System Sample A(μmol / L) B (Salt w / Spiked w / (Originalin Model Reduced) Peptide) Peptide)Formulation)Broth47.7 n / a95.4 13.6Cheese1:6 milk (0.29x TC) 66.68 63.73 65.53 66.72 sauce190.8 27.3(0.57x TC)47.7 n / aCreamNone 95.4 n / a 66.26 48.42 55.58 53.24 cheese190.8 n / a47.7 n / a95.4 3.2Soy 1:29(0.07x TC) 55.16 40.49 45.47 50.53 sauce water190.8 6.4(0.13x TC)47.7 n / a95.4 13.6Tomato 1:6(0.29x TC) 42.49 46.3 49.68 50.71 ketchup water190.8 27.3(0.57x TC)47.7 n / aPlant-based 1:3 95.4 23.9(0.5x TC) 60.82 54.52 53.98 54.70 cream water190.8 47.7(Ix TC)47.7 n / a95.4 23.9Tomato 1:3(0.5x TC) 57.23 60.53 54.62 56.58 frito water190.8 47.7(lx TC)47.7 n / a95.4 47.7Turkey 1:1(lx TC) 65.93 56.50 55.33 62.55 Gravy water190.8 95.4(2x TC)47.7 n / a1:1 in 95.4 47.7Chickenchicken (Ix TC) 49.53 40.68 46.94 43.60 brothbroth 190.8 95.442Attorney Docket 1410-163200-PC

[0126] It was surprisingly found that the salty-enhancing peptides may enhance salty taste at levels in many food systems even below their salty taste threshold concentrations in the model broth. For reference, the EIV peptide had a modulating taste threshold (salty) in model broth of 20.6 μmol / L and an intrinsic taste threshold in water of 56.6 μmol / L (bitter taste); the HQPHQPLPP peptide (SEQ ID NO. 8) had a modulating taste threshold (salty) in model broth of 17.3 μmol / L and an intrinsic taste threshold in water of 119.8 μmol / L (sour taste; and the SKVLPVPQ peptide (SEQ ID NO. 12) had a modulating taste threshold (salty) in model broth of 47.7 μmol / L and an intrinsic taste threshold in water of 95.4 μmol / L (bitter), as determined in Example I.

[0127] Seven of the eight model food systems were diluted. The model food systems were prepared per the '’ full strength" formulations, spiked at 2x or 4x taste threshold, and then diluted at the ratios listed in Tables 6-8 before tasting. Because the dilutions were prepared after spiking with peptides, in some cases the peptide levels were reduced to below their taste thresholds in model broth.

[0128] Cheese Sauce. The data demonstrated that the EIV peptide may enable 20% sodium reduction in the cheese sauce when added at 2–4x TC (or 0.29–0.57x TC per the dilution). The data also demonstrated that the HQPHQPLPP peptide (SEQ ID NO. 8) may enable 20% sodium reduction in the cheese sauce when added at 4x. TC (or 0.57x TC, per the dilution). The data demonstrated that the SKVLPVPQ peptide (SEQ ID NO. 12) may enable 20% sodium reduction in the cheese sauce when added at 4x TC (or 0.57x TC, per the dilution), but there was only a minor difference between the salt intensity of the "original" and "salt-reduced" formulations.

[0129] Cream Cheese Products. The EIV peptide and HQPHQPLPP peptide (SEQ ID NO. 8) may enable 20% sodium reduction at less than 2x TC in cream cheese type products. The SKVLPVPQ peptide (SEQ ID NO. 12) provided its highest salty enhancement at the 2x TC but may enable only about 10% sodium reduction in cream cheese type products.

[0130] Soy Sauce. The EIV peptide at 2x TC (or 0.07x TC, per the dilution) appears to be salty enhancing. The HQPHQPLPP peptide (SEQ ID NO. 8) at 2x TC (or 0.07x TC, per the dilution) appears to be salty enhancing but with no greater benefit above the 2x threshold 43Attorney Docket 1410-163200-PCconcentration. The SKVLPVPQ peptide (SEQ ID NO. 12) appears to be salty enhancing at increasing levels of the peptide. However, because soy sauce has a strong umami taste and saltiness was hard to distinguish, the evaluation of the peptide in soy sauce may be less reliable than the other food systems.

[0131] Tomato Ketchup. The EIV peptide at 2x TC (or 0.29x TC, per the dilution) appears to be salty enhancing with no greater benefit at 4x TC. The EIV peptide may enable 20% sodium reduction at less than 2x TC. The HQPHQPLPP peptide (SEQ ID NO. 8) at 4x TC (or 0.57x TC. per the dilution) appears to be salty enhancing. The SKVLPVPQ peptide (SEQ ID NO.12) appears to be salty enhancing at increasing levels of the peptide. However, the performance of the peptides in the tomato ketchup was difficult to evaluate due to strong sour, salty, and sweet taste intensities.

[0132] Plant-Based Cream Cheese Spread. None of the EIV peptide, the HQPHQPLPP peptide (SEQ ID NO. 8), and the SKVLPVPQ peptide (SEQ ID NO. 12) provided a prominent salty enhancing effect in the plant-based cream cheese product. Interactions between plant proteins and peptides may play a role. It is possible that higher dosages of the peptides may be helpful.

[0133] Tomato Frito. The EIV peptide and HQPHQPLPP peptide (SEQ ID NO. 8) may enable 20% sodium reduction at about 2x TC (or 0.5x TC, per the dilution). The SKVLPVPQ peptide (SEQ ID NO. 12) was not taste modulating in this food matrix at the evaluated concentrations.

[0134] Turkey Gravy. The EIV peptide and HQPHQPLPP peptide (SEQ ID NO.8) were found not to be taste modulating in the turkey gravy system at the evaluated concentrations. The SKVLPVPQ peptide (SEQ ID NO. 12) may enable 20% sodium reduction at greater than 4x TC or enable about 15 % sodium reduction at 4x TC (or 2x TC, per the dilution).

[0135] Chicken Broth. The EIV peptide may enable 20% sodium reduction in this type of food matrix at 2x TC (or 1 x TC, per the dilution) but had only a minor different between the "original" formulation of the broth and the salt-reduced broth. The HQPHQPLPP peptide (SEQ ID NO. 8) may enable greater than 20% sodium reduction at 2x TC (or lx TC, per the dilution)44Attorney Docket 1410-163200-PCin this type of food matrix. The SKVLPVPQ peptide (SEQ ID NO. 12) contributed its greatest salty enhancement at 2x TC (or lx TC, per the dilution) but may enable only about 10% sodium reduction on its own in this type of food matrix.

[0136] Overall, it was unexpected that the SKVLPVPQ peptide (SEQ ID NO. 12) behaved differently from the other two peptides, particularly because the SKVLPVPQ peptide (SEQ ID NO. 12) had a similar length and amino acids as in HQPHQPLPP (SEQ ID NO. 8). Each of the peptides performed in a way that was unexpected based on its amino acid composition and length,

[0137] No trends were identified in how dilution impacts peptide performance. All three peptides were effective at low or no dilution (i.e., 1:1 dilution or undiluted) to higher dilutions (e.g., 1:6 or 1:29 dilution).

[0138] Further, no trends were identified in how the ingredient characteristics of the diluted model systems (e.g., moisture, fat, pH, salt, dairy vs non-dairy) influence peptide performance at the levels tested in the model food systems. Similarly, no trends were identified in the ingredient characteristics of undiluted model systems as influencing peptide performance at the levels tested in the model food systems. It is possible that other components in the model food systems (e.g., presence of glutamic acid, aspartic acid, amino acids, salt, and / or organic acids) may impact how the peptides perform.

[0139] All percentages described herein are percent by weight unless otherwise specified. The description herein is not to be taken in a limiting sense but is made merely for the purpose of describing the general principles of exemplary embodiments. Reference throughout this specification to ''one embodiment," "an embodiment," "some embodiments", "an implementation", "some implementations", "some applications", or similar language means that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one embodiment of the present invention. Thus, appearances of the phrases "in one embodiment," "in an embodiment," "in some embodiments", "in some implementations", and similar language throughout this specification may, but do not necessarily, all refer to the same embodiment.

[0140] Those skilled in the art will recognize that a wide variety of other modifications, alterations, and combinations can also be made with respect to the above-described embodiments without departing from the scope of the invention, and that such modifications, alterations, and combinations are to be viewed as being within the ambit of the inventive concept.

Claims

CLAIMSWhat is claimed is:

1. A taste modifier composition comprising:a food grade carrier; andone or more of the following salty-enhancing peptides:EIV peptide in an amount of at least 0.1 wt.%; and / orHQPHQPLPP peptide (SEQ ID NO. 8) in an amount of at least 0.1 wt.%; and / or SKVLPVPQ peptide (SEQ ID NO. 12) in an amount of at least 0.1 wt.%.

2. The taste modifier composition according to claim 1, wherein the one or more salty- enhancing peptides comprises the EIV peptide.

3. The taste modifier composition according to claim 1, wherein the one or more salty-enhancing peptides comprises the HQPHQPLPP peptide (SEQ ID NO. 8).

4. The taste modifier composition according to claim 1, wherein the one or more salty- enhancing peptides comprises the SKVLPVPQ peptide (SEQ ID NO. 12).

5. The taste modifier composition according to claim 1, wherein the salty-enhancing peptides comprise one or more of:(i) the EIV peptide and the HQPHQPLPP peptide (SEQ ID NO. 8);(ii) the EIV peptide and the SKVLPVPQ peptide (SEQ ID NO. 12);(iii) the HQPHQPLPP peptide (SEQ ID NO. 8) and the SKVLPVPQ peptide (SEQ ID NO.12); or(iv) the EIV peptide, the HQPHQPLPP peptide (SEQ ID NO. 8), and the SKVLPVPQ peptide (SEQ ID NO. 12).

6. The taste modifier composition according to any one of claims 1 to 5. comprising one or more of:the EIV peptide in an amount of at least 2 wt.%; and / or47Attorney Docket 1410-163200-PCthe HQPHQPLPP peptide (SEQ ID NO. 8) in an amount of at least 2 wt.%; and / or the SKVLPVPQ peptide (SEQ ID NO. 12) in an amount of at least 2 wt.%.

7. The taste modifier composition according to any one of claims 1 to 6, wherein the food grade carrier is one or more of water, propylene glycol, 1,3-propanediol, ethanol, glycerin, and lactic acid and the taste modifier composition is in liquid form.

8. The taste modifier composition according to any one of claims 1 to 6, wherein the food grade carrier comprises one or more of gum arabic, dextrose, sucrose, fructose, lactose, protein, cellulose, silica, calcium silicate, tapioca flour, rice flour, salt, potassium chloride, starch, and / or maltodextrin, and the taste modifier composition is in powder or particulate form.

9. The taste modifier composition according to any one of claims 1 to 6, wherein the taste modifier composition is in the form of a cultured milk or culture medium.

10. A salty taste-enhanced food or beverage product comprising the taste modifier composition of claim 1 as an added ingredient of the salty taste-enhanced food or beverage product, and the taste modifier composition is included in an amount effective to provide at least one of the salty -enhancing peptides in one or more of the following amounts:the EIV peptide in an amount of at least 0.000175% by weight of the food or beverage product; and / orthe HQPHQPLPP peptide (SEQ ID NO. 8) in an amount of at least 0.00045% by weight of the food or beverage product; and / orthe SKVLPVPQ peptide (SEQ ID NO. 12) in an amount of at least 0.001025% by weight of the food or beverage product.

11. The salty taste-enhanced food or beverage product according to claim 10, wherein the taste modifier composition is included in an amount effective to provide at least one of the salty-enhancing peptides in one or more of the following amounts:the EIV peptide in an amount of at least 0.0007% by weight of the food or beverage product; and / or48Attorney Docket 1410-163200-PCthe HQPHQPLPP peptide (SEQ ID NO. 8) in an amount of at least 0.0018% by weight of the food or beverage product; and / orthe SKVLPVPQ peptide (SEQ ID NO. 12) in an amount of al least 0.0041% by weight of the food or beverage product.

12. The salty taste-enhanced food or beverage product according to claim 10, wherein: the EIV peptide is included in an amount of at least 0.000175% to less than 0.00203% by weight of the food or beverage product; and / orthe HQPHQPLPP peptide (SEQ ID NO. 8) in an amount of at least 0.00045% to less than 0.01258% by weight of the food or beverage product; and / orthe SKVLPVPQ peptide (SEQ ID NO. 12) in an amount of at least 0.001025% to less than 0.00827% by weight of the food or beverage product.

13. The salty taste-enhanced food or beverage product according to claim 10 or 12, wherein the salty taste-enhanced food is a dairy-based or plant-based food product selected from process cheese, cream cheese, cheese powder, cheese sauce, sauce, condiment, broth, and gravy.

14. A method of enhancing a salty taste of a food or beverage product, the method comprising adding to the food or beverage product one or more salty-enhancing peptides: the EIV peptide in an amount of at least 0.000175% by weight of the food or beverage product; and / orthe HQPHQPLPP peptide (SEQ ID NO. 8) in an amount of at least 0.00045% by weight of the food or beverage product; and / orthe SKVLPVPQ peptide (SEQ ID NO. 12) in an amount of at least 0.001025% by weight of the food or beverage product.

15. The method according to claim 14, wherein the one or more salty-enhancing peptides comprises the EIV peptide.

16. The method according to claim 14, wherein the one or more salty-enhancing peptides comprises the HQPHQPLPP peptide (SEQ ID NO. 8).49Attorney Docket 1410-163200-PC17. The method according to claim 14, wherein the one or more salty -enhancing peptides comprises the SKVLPVPQ peptide (SEQ ID NO. 12).

18. The method according to any one of claims 14 to 17, wherein the food product is a dairy¬ based or plant-based food product selected from process cheese, cream cheese, cheese powder, cheese sauce, sauce, condiment, broth, and gravy.

19. The method according to any one of claims 14 to 19, wherein the food or beverage product includes at least one of the salty-enhancing peptides in one or more of the following amounts:EIV peptide in an amount of at least 0.0007% by weight of the food or beverage product; and / orHQPHQPLPP peptide (SEQ ID NO. 8) in an amount of at least 0.0018% by weight of the food or beverage product; and / orSKVLPVPQ peptide (SEQ ID NO. 12) in an amount of at least 0.0041% by weight of the food or beverage product.

20. The method according to any one of claims 14 to 18, wherein:the EIV peptide is included in an amount of at least 0.000175% to less than 0.00203% by weight of the food or beverage product; and / orthe HQPHQPLPP peptide (SEQ ID NO. 8) in an amount of at least 0.00045% to less than 0.01258% by weight of the food or beverage product; and / orthe SKVLPVPQ peptide (SEQ ID NO. 12) in an amount of at least 0.001025% to less than 0.00827% by weight of the food or beverage product.

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