Method for producing fermented dairy product with improved stability

Abstract

The disclosure belongs to the technical field of dairy products. The present disclosure relates to methods of producing acidified or fermented dairy products with improved stability. The present disclosure also provides acidified or fermented dairy products produced therefrom.

Description

[0001] field

[0002] The present disclosure generally relates to the field of acidified or fermented milk, and more particularly to a method for producing an acidified or fermented milk product with improved stability, and an acidified or fermented milk product prepared by the method.

[0003] background

[0004] Flavor and gas formation are important attributes that influence consumer acceptance of acidified or fermented dairy products. Fermented dairy products can experience changes in flavor and / or gas formation due to a lack of cooling or temperature fluctuations during transportation and storage. In addition to changes in the product's taste and mouthfeel, gas formation can also lead to increased pressure, expansion, and potential bursting of finished packaging.

[0005] Therefore, there is a need for improved methods of producing acidified or fermented dairy products that remain stable during storage, particularly when a cold chain is not possible or is at risk of being damaged or partially interrupted.

[0006] Overview

[0007] In a first aspect, the present disclosure provides a method for producing an acidified or fermented dairy product comprising the steps of:

[0008] (a) adding one or more sweeteners to a dairy base or pre-acidified milk, wherein the dairy base is acidified until a first target pH of not more than 4.7 is reached, to obtain pre-acidified milk;

[0009] (b) optionally heat-treating the pre-acidified milk;

[0010] (c) inoculating the pre-acidified milk with a lactose-deficient lactic acid bacterium until a second target pH of no more than 4.6 is reached to obtain a dairy product; and

[0011] wherein conversion of citric acid to acetic acid in the dairy product is delayed during storage compared to a dairy product produced without the one or more sweeteners.

[0012] In a second embodiment, the present disclosure provides an acidified or fermented dairy product obtained by the present method. Detailed Description of the Invention

[0014] As described below, it has surprisingly been found that by delaying the conversion of citric acid to acetic acid, off-flavor and gas formation in acidified or fermented dairy products can be controlled during shelf life. The present disclosure provides a solution to controlling the conversion by adding specific sweeteners as described in more detail above.

[0015] The present disclosure relates to a method for producing an acidified or fermented dairy product comprising the following steps:

[0016] (a) adding one or more sweeteners to a dairy base or pre-acidified milk, wherein the dairy base is acidified until a first target pH of not more than 4.7 is reached, to obtain pre-acidified milk;

[0017] (b) optionally heat-treating the pre-acidified milk;

[0018] (c) inoculating the pre-acidified milk with a lactose-deficient lactic acid bacterium until a second target pH of no more than 4.6 is reached to obtain an acidified dairy product; and

[0019] wherein conversion of citric acid to acetic acid in the dairy product during storage is delayed compared to an acidified dairy product produced without the use of the one or more sweeteners.

[0020] In one embodiment, the present disclosure relates to a method wherein the lactose deficient lactic acid bacterium in step (c) is a strain of the genus Lactobacillus.

[0021] In one embodiment, the present disclosure relates to a method wherein the strain of the genus Lactobacillus is selected from the species Lacticaseibacillus rhamnosus, Lacticaseibacillus casei, and Lacticaseibacillus paracasei.

[0022] In one embodiment, the present disclosure relates to a method wherein the lactose-deficient lactic acid bacteria strain in step (c) is one or more selected from ATCC53103, CNCM 1-2116 and DSM16572.

[0023] In one embodiment, the present disclosure relates to a method wherein the lactose-deficient lactic acid bacteria in step (c) ferment the pre-acidified milk to obtain a fermented dairy product.

[0024] The term "milk" is to be understood as the lacteal secretion obtained by milking an animal, such as any mammal, including but not limited to cows, sheep, goats, buffaloes, camels, llamas, mares and deer. In a preferred embodiment, the milk is bovine milk.

[0025] A "milk base" can be any raw milk and / or processed milk component or other material derived from milk that can be acidified according to the method of the present invention. Thus, useful milk bases include (but are not limited to) solutions / suspensions of any milk or milk-like product containing proteins, such as whole milk, whole milk, fat-free milk, low-fat milk, skim milk, buttermilk, lactose-reduced milk, concentrated milk, reconstituted milk powder, condensed milk, dry milk, whey, whey permeate, lactose, mother liquor obtained by crystallization of lactose, whey protein concentrate or cream. Obviously, the milk base can be derived from any mammal, such as essentially pure mammalian milk, or reconstituted milk powder. Preferably, at least part of the protein in the milk base is a protein naturally occurring in mammalian milk, such as casein or whey protein.

[0026] In one embodiment, the present disclosure relates to a method wherein the milk base is derived from an animal, such as a mammal. In one embodiment, the present disclosure relates to a method wherein the mammal is selected from the group consisting of a cow, sheep, goat, buffalo, camel, llama, mare, and deer. In a preferred embodiment, the mammal is a cow.

[0027] Prior to acidification, the milk base may be homogenized and pasteurized according to methods known in the art. "Homogenization" as used herein means mixing thoroughly to obtain a soluble suspension or emulsion. If homogenization is performed before fermentation, this can break down the milk fat into smaller sizes so that it can no longer separate from the milk. This can be achieved by forcing the milk through small holes under high pressure. "Pasteurization" as used herein means treating the milk base to reduce or eliminate the presence of living organisms, such as microorganisms. Preferably, pasteurization is achieved by maintaining a specified temperature for a specified period of time. The specified temperature is generally achieved by heating. The temperature and duration can be selected so as to kill or inactivate specific bacteria, such as harmful bacteria. A rapid cooling step may then be performed.

[0028] Milk bases derived from mammals contain lactose as the main carbohydrate. Lactose is hydrolyzed into the monosaccharides glucose and galactose by lactic acid bacteria that metabolize lactose (lactose positive). If the lactic acid bacteria cannot metabolize lactose, i.e., are lactose deficient, or if the pre-acidification of the milk base is chemical acidification, it may be necessary to add a suitable carbohydrate to the milk base or the pre-acidified milk to obtain at least one carbohydrate and / or monosaccharide that the lactic acid bacteria can use to ferment to the second target pH.

[0029] In the context of the present invention, the term "lactose-deficient" is used to characterize lactic acid bacteria that have partially or completely lost the ability to use lactose as a source of cell viability or growth. Lactose-deficient bacteria are able to metabolize one or more carbohydrates selected from sucrose, galactose, glucose, and / or other fermentable carbohydrates. Since these carbohydrates are not naturally present in milk in sufficient quantities to support fermentation by lactose-deficient bacteria, they must be added to the milk base or pre-acidified milk.

[0030] When the first target pH is reached, pre-acidification is terminated. The first target pH must be equal to or higher than the second target pH and can be selected to provide a pH range that allows for potential further acidification to the second target pH. In one embodiment, the present disclosure relates to a method wherein the first target pH is no greater than pH 4.70: is pH 4.65, pH 4.60, pH 4.55, pH 4.50, pH 4.45, pH 4.40; or is in the range of pH 4.70-4.00, pH 4.70-4.10, pH 4.70-4.20, pH 4.70-4.30, pH 4.70-4.40, pH 4.70-4.45, pH 4.65-4.50, pH 4.60-4.55; or is about 4.70, 4.65, 4.60, 4.55, 4.50, 4.45, or 4.40.

[0031] In one embodiment, the present disclosure relates to fermentations wherein the second target pH is no more than pH 4.5, pH 4.4, pH 4.3, pH 4.2, pH 4.1, pH 4.0, pH 3.9, pH 3.8, pH 3.7, pH 3.6, pH 3.5; or in the range of pH 4.50-3.50, pH 4.50-4.05, pH 4.45-4.10, pH 4.45-4.15, pH 4.40-4.20, pH 4.40-4.25, pH 4.35-4.30, pH 4.00-3.50, pH 3.95-3.50, pH 3.90-3.55, pH 3.85-3.60, pH 3.80-3.65, pH 3.75-3.60; or about pH 4.40, pH 4.35, pH 4.30, pH 4.25, pH 4.20, pH 4.15, pH 4.10, pH 4.05, pH 4.00, pH 3.90, pH 3.80, pH 3.70, pH 3.60, pH 3.50.

[0032] Temperature affects the rate of acidification and fermentation and is preferably kept stable or constant at a specified temperature during the acidification process. In one embodiment, the present disclosure relates to a method wherein the acidification and / or fermentation temperature does not exceed 25°C, 30°C, 35°C, 36°C, 37°C, 38°C, 39°C, 40°C, 41°C, 42°C, 43°C, 44°C, 45°C; or is in the range of 20°C-45°C, 25°C-45°C, 30°C-45°C, 40°C-45°C, 25°C-40°C, 30°C-40°C, 35°C-40°C; or is about 20°C, 25°C, 30°C, 35°C, 36°C, 37°C, 38°C, 39°C, 40°C, 41°C, 42°C, 43°C, 44°C, 45°C.

[0033] In one embodiment, the present disclosure relates to methods wherein the one or more sweeteners is a sugar and / or a sugar alcohol.

[0034] In one embodiment, the present disclosure relates to a method wherein the sugar is selected from fructose, galactose, glucose, and sucrose. In one embodiment, the present disclosure relates to a method wherein the concentration of the sugar is 0.001%, 0.005%, 0.01%, 0.05%, 0.10%, 0.15%, 0.20%, 0.25%, 0.30%, 0.35%, 0.40%, 0.45%, or 0.50%, or within the range of 0.05%-0.50%, 0.10%-0.40%, or 0.15%-0.30%. Unless otherwise indicated, percentages (%) in the present disclosure are expressed as weight / volume (w / v), i.e., % w / v.

[0035] In one embodiment, the present disclosure relates to the method wherein the sugar is fructose at a concentration of 0.001%, 0.005%, 0.01%, 0.05%, 0.10%, 0.15%, 0.20%, 0.25%, or in the range of 0.001%-0.10%, 0.10%-0.25%, 0.15%-0.20%. In one embodiment, the present disclosure relates to the method wherein the sugar is glucose at a concentration of 0.001%, 0.005%, 0.01%, 0.05%, 0.10%, 0.15%, 0.20%, 0.25%, or in the range of 0.001%-0.10%, 0.10%-0.25%, 0.15%-0.20%. In one embodiment, the present disclosure relates to the method, wherein the sugar is a combination of fructose and glucose, wherein the concentration of fructose is 0.001%, 0.005%, 0.01%, 0.05%, 0.10%, 0.15%, 0.20%, 0.25% or within the range of 0.001%-0.10%, 0.10-0.25%, 0.15-0.20% and the concentration of glucose is 0.001%, 0.005%, 0.01%, 0.05%, 0.10%, 0.15%, 0.20%, 0.25% or within the range of 0.001%-0.10%, 0.10-0.25%, 0.15-0.20%. In one embodiment, the present disclosure relates to the method wherein the sugar is 0.10% fructose and 0.10% glucose, 0.15% fructose and 0.15% glucose, 0.20% fructose and 0.20% glucose, 0.10% fructose and 0.15% glucose, 0.10% fructose and 0.20% glucose, 0.15% fructose and 0.10% glucose, or 0.20% fructose and 0.10% glucose.

[0036] In one embodiment, the present disclosure relates to the method, wherein the one or more sweeteners are selected from C4 sugar alcohols [C4H 10 O4] and C5 sugar alcohols [C5H 12 O5] sugar alcohol.

[0037] In one embodiment, the present disclosure relates to the method, wherein the C4 sugar alcohol is erythritol, D-threitol or L-threitol; and the C5 sugar alcohol is xylitol, ribitol, D-arabitol, L-arabitol, D-lyxitol or L-lyxitol. In one embodiment, the present disclosure relates to the method, wherein the concentration of the sugar alcohol is 0.001%, 0.005%, 0.01%, 0.05%, 0.10%, 0.15%, 0.20%, 0.25%, 0.5%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5% or 5.0%, or in the range of 0.5%-5.0%, 1.0%-4.5%, 1.5%-4.0%, 2.0%-3.5% or 2.5%-3.0%.

[0038] In one embodiment, the present disclosure relates to the method, wherein the sugar alcohol is xylitol and its concentration is selected from 0.001%, 0.005%, 0.01%, 0.05%, 0.10%, 0.15%, 0.20%, 0.25%, 0.5%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5% or 5.0%, or is within the range of 0.5%-5.0%, 1.0%-4.5%, 1.5%-4.0%, 2.0%-3.5% or 2.5%-3.0%. In one embodiment, the present disclosure relates to the method, wherein the sugar alcohol is xylitol at a concentration of 2.0%.

[0039] Sugar alcohol can be added as a single sugar alcohol or as a mixture of two or more sugar alcohols. In one embodiment, the present disclosure relates to a combination of at least one C4 sugar alcohol and at least one C5 sugar alcohol. In one embodiment, at least one C4 sugar alcohol comprises erythritol. In one embodiment, at least one C5 sugar alcohol comprises xylitol. In one embodiment, the present disclosure relates to a combination comprising or consisting of erythritol and xylitol.

[0040] The ratio of C4:C5 sugar alcohols can vary. In one embodiment, the ratio of C4:C5 is selected from 1:99, 10:90, 20:80, 30:70, 40:60, 45:55, 50:50, 55:45, 60:40, 70:30, 80:20, 90:10, or 99:1. In one embodiment, the ratio of erythritol to xylitol (Ery:Xyl) is selected from 1:99, 10:90, 20:80, 30:70, 40:60, 45:55, 50:50, 55:45, 60:40, 70:30, 80:20, 90:10, or 99:1.

[0041] In one embodiment, the present disclosure relates to the method, wherein the one or more sweeteners are a combination of a sugar alcohol and a sugar. In one embodiment, the present disclosure relates to the method, wherein the one or more sweeteners comprise a combination of xylitol and fructose and / or glucose. In one embodiment, the present disclosure relates to the method, wherein the one or more sweeteners comprise a combination of 2% xylitol and 0.1%-0.2% fructose and / or 0.1%-0.2% glucose.

[0042] In one embodiment, the present disclosure relates to the method wherein the one or more sweeteners are added to the milk base. In one embodiment, the present disclosure relates to the method wherein the one or more sweeteners are added to pre-acidified milk. In one embodiment, the present disclosure relates to the method wherein the one or more sweeteners are added to heat-treated and pre-acidified milk.

[0043] As will be apparent from this disclosure, by carefully selecting the type, concentration, and combination of one or more sweeteners, it is possible to control the conversion of citric acid to acetic acid, thereby providing a method for producing a dairy product having improved stability in terms of flavor, gas formation, taste, and mouthfeel. In one embodiment, the present disclosure relates to controlling the conversion of citric acid to acetic acid using one or more sweeteners. The specific combinations disclosed in the methods and examples also apply to controlling the conversion using one or more sweeteners.

[0044] Acidification can be achieved by chemical acidification by adding an acid or an acid-containing composition. Examples of acids include, but are not limited to, acetic acid (common in vinegar), adipic acid, citric acid (common in lemon juice), fumaric acid, glucono-δ-lactone, hydrochloric acid, lactic acid, malic acid, phosphoric acid, succinic acid, and tartaric acid. In one embodiment, the present disclosure relates to the method wherein chemical acidification is due to the addition of an acid selected from the group consisting of acetic acid, adipic acid, citric acid, fumaric acid, glucono-δ-lactone, hydrochloric acid, lactic acid, malic acid, phosphoric acid, succinic acid, and tartaric acid.

[0045] Acidification may also be a result of fermentation by lactic acid bacteria when lactic acid bacteria are added to a milk base or pre-acidified milk comprising one or more substrates fermentable by lactic acid bacteria.

[0046] Acidification may also be the result of a combination of chemical acidification and fermentation by lactic acid bacteria. Chemical acidification and fermentation by lactic acid bacteria can be performed in the same step or in different steps. In one embodiment, the disclosure relates to a method wherein acidification is the result of both chemical acidification and fermentation by lactic acid bacteria.

[0047] In one embodiment, the present disclosure relates to the method, wherein the pre-acidification of the milk base in step (a) is chemical acidification and / or due to fermentation by one or more lactic acid bacteria.

[0048] In one embodiment, the present disclosure relates to the process, wherein the pre-acidification of the milk base in step (a) is due to fermentation by one or more lactic acid bacteria, and wherein the pre-acidified milk is heat treated.

[0049] In one embodiment, the present disclosure relates to the method, wherein the one or more lactic acid bacteria belong to the genus Streptococcus, such as S. thermophilus, or to the genus Lactobacillus, such as L. delbrueckii subsp. Bulgaricus.

[0050] One or more lactic acid bacteria used to acidify the milk base to obtain pre-acidified milk may be lactose-deficient. Examples of suitable lactose-deficient strains can be found in WO2015 / 193459. In one embodiment, the present disclosure relates to a method in which one or more lactic acid bacteria are lactose-deficient. In one embodiment, the present invention relates to a method in which the lactose-deficient strain is selected from the group consisting of DSM28952, DSM28953, DSM28910, DSM32600 and DSM32599.

[0051] In a second aspect, the present disclosure relates to an acidified or fermented dairy product obtained by the method described herein. Therefore, any disclosure made on the method may also apply to the dairy product obtained. As used herein, the term "acidified dairy product" or "fermented dairy product" refers to a food or feed product, wherein the preparation of the food or feed product involves acidification and / or fermentation of a milk base (see above). According to the present disclosure, the fermentation is carried out by lactic acid bacteria. "Acidified dairy product" and "fermented dairy product" as used herein include but are not limited to dairy products, such as yogurt. In one embodiment, the present disclosure relates to an acidified or fermented dairy product, which is a food or feed product. In one embodiment, the present disclosure relates to an acidified or fermented dairy product, which is a dairy product, such as yogurt (set or stirred); Greek yogurt; yogurt-based products such as fruit yogurt, and yogurt-based beverages such as drinking yogurt; buttermilk; kefir; labneh, quark. Preferably, the acidified or fermented dairy product is yogurt.

[0052] In the context herein, the term "starter" is a culture, which is a preparation (composition) of one or more bacterial strains (such as lactic acid bacteria) to assist the start of a fermentation process in the preparation of fermented products such as various foods, feeds and beverages. In this context, a "yogurt starter" is a bacterial culture comprising one or more lactobacillus bulgaricus strains and / or one or more thermophilus streptococci strains. Accordingly, "yogurt" refers to a fermented dairy product that can be obtained by inoculating and fermenting a milk base with a composition comprising lactobacillus bulgaricus and thermophilus streptococci strains.

[0053] In one embodiment, the present disclosure relates to an acidified or fermented dairy product, wherein the product is an ambient storage product. The term "ambient storage product" or "ambient storage yogurt" refers to a product suitable for storage at ambient temperature for a period of time. The storage period can be between 1 month and 12 months, for example 1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months or 12 months. The term "ambient temperature" or alternatively "room temperature" in this context refers to a temperature above 10°C, 15°C, 20°C, 25°C or between 10°C-50°C, 10°C-40°C, 10°C-30°C, 15°C-45°C, 15°C-35°C, 15°C-25°C, 20°C-40°C or 20°C-30°C.

[0054] In one embodiment, the present disclosure relates to an acidified or fermented dairy product, wherein the product is a pasteurized product, such as pasteurized yogurt (PPY). The term "pasteurized product" or "pasteurized yogurt" refers to a product that has been heat treated (pasteurized) after the acidification / fermentation step.

[0055] Acidified or fermented dairy products typically contain protein at a level of 2.0% to 3.5% w / w. Acidified or fermented dairy products can be low-protein products with a protein level between 1.0% and 2.0% w / w. Alternatively, acidified or fermented dairy products can be high-protein products with a protein level greater than 3.5% or 5.1% w / w, for example, between 3.5% and 5.1%, 3.5% and 10.5%, or 5.1% and 10.5% w / w. The protein can be derived from milk, such as whey or casein.

[0056] The product obtained by the method of the present disclosure may include additional ingredients. For example, if the sweetness is not high enough, additional sweeteners such as artificial sweeteners such as aspartame, acesulfame potassium (Ace-K), sucralose, neotame, advantame and saccharin, or sweet mixtures such as preparations containing fruit can be added. In one embodiment of the present disclosure, the product further includes an additional sweetening ingredient. In one embodiment, the sweetening ingredient is an artificial sweetener or a sweet mixture. In one embodiment, the sweetener is selected from aspartame, acesulfame potassium (Ace-K), sucralose, neotame, advantame and saccharin. In one embodiment, the sweet mixture is a preparation containing fruit.

[0057] The method for producing a fermented product described in the present disclosure is suitable for producing a product containing probiotic lactic acid bacteria.

[0058] In one embodiment, the present disclosure relates to the method, wherein the one or more lactic acid bacteria of the second culture are selected from the group consisting of: bacteria of the genus Lactobacillus, such as Lactobacillus acidophilus, Lacticaseibacillus paracasei, Lacticaseibacillus rhamnosus, Lacticaseibacillus casei, Lactobacillus delbrueckii, Lactiplantibacillus plantarum, Limosilactobacillus fermentum, Limosilactobacillus reuteri, and Lactobacillus johnsonii; bacteria of the genus Bifidobacterium, such as Bifidobacterium longum, Bifidobacterium adolescentis, and Lactobacillus johnsonii. adolescentis), Bifidobacterium bifidum, Bifidobacterium breve, Bifidobacterium animalis subsp. lactis, Bifidobacterium dentium, Bifidobacterium catenulatum, Bifidobacterium angulatum, Bifidobacterium magnum, Bifidobacterium pseudocatenulatum and Bifidobacterium infantis; or Streptococcus, such as S. thermophilus.

[0059] Consumption of probiotics is considered to be beneficial to an individual's health. Therefore, a certain amount of live probiotics is required in fermented dairy products. In one embodiment, the present invention relates to a method wherein the one or more lactic acid bacteria of the second culture are probiotics.

[0060] In particular, probiotic lactic acid bacteria are lactose deficient (lactose negative). The effectiveness of probiotic lactic acid bacteria depends on the number of cells present in the fermented product, i.e. the cell count. The cell count is a number given as colony forming units per gram of fermented product, i.e. cfu / g. In one embodiment of the present disclosure, the amount of probiotic lactic acid bacteria in the fermented product after storage at ambient temperature for 3 months is greater than 1E+05 cfu / g, greater than 5E+05 cfU / g, greater than 1E+06 cfu / g, greater than 5E+06 cfU / g, greater than 1E+07 cfu / g, greater than 5E+07 cfU / g, greater than 1E+08 cfu / g, greater than 5E+08 cfu / g, greater than 1E+09 cfu / g, greater than 5E+09 cfu / g, greater than 1E+10 cfu / g, greater than 5E+10 cfu / g, greater than 1E+11 cfu / g, greater than 5E+11 cfu / g, greater than 1E+12 cfu / g, greater than 5E+12 cfu / g. In one embodiment, the ambient temperature is 25° C. In another embodiment, the ambient temperature is 37° C. In a preferred embodiment, the amount of lactic acid bacteria is greater than 5E+06 cfu / g after storage for 6 months at 25° C. In another preferred embodiment, the amount of lactic acid bacteria is greater than 5E+06 cfu / g after storage for 4 months at 37° C. Example

[0061] Example 1 - comprising Citric acid consumption and acetic acid production during the shelf life of pasteurized yogurt.

[0062] The results showed that the growth of Lactobacillus rhamnosus strain (F-DVS) in pasteurized yogurt (PPY) during the shelf life was 1. Hansen) to convert citric acid into acetic acid.

[0063] Briefly, PPY was produced as follows. A milk base (containing 3.5% of the sugar alcohol maltitol ( P200, Roquette) and 1% erythritol (Zerose TM The milk was heat treated at 134°C for 4 seconds using erythritol 16952 (Cargill) and then cooled to 5°C. The milk base was fermented at 43°C to a first target pH of 4.45 using F-DVS Acidifix 1.0 (Chr. Hansen). The fermented milk was heat treated at 75°C for 20 seconds and aseptically filled in cups and refrigerated at 6°C. For the second fermentation, F-DVS Hansen) was diluted in milk B (reconstituted skim milk containing 9.5% MSNF, heat-treated at 99°C for 30 min) and inoculated at 1 mL / 100 mL cup to obtain a final concentration of 5.3E+06 CFU / g yogurt. Fermentation was carried out at 25°C for 72 hours to reach a second target pH of ≤4.41. No citric acid consumption was observed during the second fermentation. Samples were stored at 25°C or 37°C. The concentrations of citric acid and acetic acid changed over time (see table below).

[0064] Table 1: Concentrations of citric acid and acetic acid (g / L) at different storage temperatures and times.

[0065] temperature Time 0 1 month 2 months 3 months 4 months 6 months citric acid 25℃ 1.30 1.33 0.59 0 0.01 0.01 Acetic acid 25℃ 0.26 0.20 0.68 1.09 1.25 1.26 citric acid 37℃ 1.30 0 NA 0 NA NA Acetic acid 37℃ 0.26 1.13 NA 1.20 NA NA

[0066] Example 2 - Erythritol and maltitol Inhibitory effect of citrate conversion.

[0067] The effects of erythritol (ERY) and / or maltitol (MAL) on the conversion of citric acid to acetic acid in pre-acidified milk supplemented with different concentrations of fructose (FRU), galactose (GAL) and / or sucrose (SUC) were investigated.

[0068] Prepare pre-acidified milk at pH 4.5 by adding 3 mL of 5 M HCl to 210 mL of milk to avoid affecting the growth period. The concentration of lactic acid produced is shown in the table below. After three days of growth in 2 mL of the different milks, the milk products were analyzed for citric acid (data not shown) and acetic acid (see table below) concentrations.

[0069] Table 2: Erythritol, but not maltitol, has an effect on acetic acid concentration (g / L).

[0070]

[0071] Example 3 - Reduced CO2 formation correlates with reduced acetic acid production.

[0072] Researched using Correlation between CO2 bubble formation, citric and acetic acid concentrations, and the presence of different concentrations of galactose and sucrose in fermented pre-acidified milk.

[0073] Prepare pre-acidified milk at pH 4.5 by adding 3 mL of 5 M HCl to 210 mL of milk. After 40 hours of growth in 2 mL of different milks, duplicates were made, after which the milks were analyzed for citric acid concentration and acetic acid content. In addition, the bottom of the plates was visually inspected four times to quantify the formation of air (CO2) bubbles.

[0074] Table 3A: Correlation of CO2 bubble formation with citric acid and acetic acid concentrations in the presence of different concentrations of galactose.

[0075] Galactose (g / L) Bubble counting Citric acid (g / L) Acetic acid (g / L) 0 10.5±1.9 0±0 0.91±0.01 0.20 4.3±1.5 0±0 0.90±0.03 0.25 3.6±1.5 0.42±0.01 0.81±0.02 0.30 3.3±1.5 0.56±0.15 0.65±0.01 0.35 1.7±0.6 0.79±0.01 0.59±0.01 0.40 0.3±0.6 0.94±0.01 0.51±0.02 0.50 0±0 1.17±0.04 0.39±0.01

[0076] Table 3B: Correlation between CO2 bubble formation and citric acid and acetic acid concentrations in the presence of different fructose concentrations.

[0077] Fructose (g / L) Bubble counting Citric acid (g / L) Acetic acid (g / L) 0 10.5±1.9 0±0 0.91±0.01 0.10 4.75±1.7 0.54±0.3 0.68±0.01 0.20 1.25±0.9 1.62±0.01 0.15±0.01 0.50 0.25±0.5 1.62±0.00 0.15±0.02

[0078] Example 4 - Available sugars and pH Effects of citric acid utilization.

[0079] We studied the effects of adding different sugars to milk. Effect of citric acid consumption. Milk and pre-acidified milk were supplemented with the following sugars: glucose (GLC), galactose (GAL), or fructose (FRU). Pre-acidified milk at pH 4.5 was prepared by adding 3 mL of 5 M HCl to 210 mL of milk. The cells were inoculated with 2 mL of different milks in 96 deep-well plates at 37°C and grown for 40 hours. The plates were stored at room temperature and the citric acid concentrations were measured by HPLC at 0, 16, 23, 40, and 500 hours. The detection limit was 0.04 g / L.

[0080] After 40 hours in milk, LGG consumed all the citric acid in the sample (Table 4A), while LGG in pre-acidified milk in the presence of low concentrations of glucose or fructose (0.1%-0.2%) resulted in no or much lower citric acid consumption (Table 4B). No added sugar and low concentrations of galactose (0.1%) also resulted in complete consumption of citric acid in pre-acidified milk. High galactose concentrations (≥0.5%) resulted in reduced consumption of citric acid in pre-acidified milk (Table 4B). This data suggests that pH and appropriate sugar availability have a significant impact on the citric acid content in milk. Citric acid consumption led to acetic acid formation in all samples (Table 4C, Table 4D).

[0081] Table 4A: Citric acid concentrations (g / L) measured during milk storage.

[0082]

[0083] Table 4B: Citric acid concentrations (g / L) measured during storage of pre-acidified milk.

[0084]

[0085] Table 4C: Acetic acid concentrations (g / L) measured during milk storage.

[0086]

[0087] Table 4D: Acetic acid concentration (g / L) measured during storage of pre-acidified milk.

[0088]

[0089] Example 5 - Sugar alcohol pairs Effects of citric acid consumption.

[0090] make The fermented milk products were grown at 37°C in pre-acidified milk supplemented with different sugars and / or sugar alcohols (at the concentrations shown in the table below). The sugars were glucose (GLC), galactose (GAL), or fructose (FRU), and the sugar alcohols were erythritol (ERY) or xylitol (XYL). The fermented milk products were stored at 37°C, and the citric acid concentration was measured after 4 days and 1 month of storage.

[0091] Surprisingly, after four days of incubation at 37°C, both erythritol and xylitol showed an effect on citric acid consumption. After one month of storage at 37°C, only xylitol and 0.2% fructose did not induce citric acid consumption. This finding suggests that citric acid consumption can be manipulated using sweeteners, namely by selecting the type and amount of sugar and / or sugar alcohol.

[0092] Table 5: Citric acid concentrations measured during storage at 37°C.

[0093]

[0094] Example 6 - Effects of sweeteners on different species of lactose-negative bacterial strains.

[0095] The effects of different sweeteners on the conversion of citric acid to acetic acid by two lactose-negative strains of Lactobacillus casei were studied.

[0096] pH 4.5 pre-acidified milk inoculated with Lactobacillus rhamnosus strain (LGG) or Lactobacillus paracasei strain (DSM16572) was prepared by adding 3ml 5M HCl to 210mL milk supplemented with different concentrations of fructose. Each bacterial strain was grown for 3 days in 2mL different milks in 96 deep-well plates at 37°C, and the content of citric acid (data not shown) and acetic acid (table 6) in the milk was analyzed afterwards. The detection limit was 0.15g / L.

[0097] Table 6: Acetic acid concentration (g / L) in milk inoculated with different lactose-negative strains

[0098]

[0099] Example 7 - Effect of adding additional sugar after the first fermentation.

[0100] The results of Example 6 indicate that the addition of additional sugar during the second fermentation delayed the conversion of citric acid to acetic acid. To further investigate the effect of adding additional sugar after the first fermentation, the following process was used. Milk bases (MB1 to MB4) were prepared and sterilized at 134°C for 4 seconds. 100 U / 1000 L F-DVS 1.0 (Chr. Hansen A / S) was inoculated into the milk base and fermented at 43°C until the first target pH of approximately 4.45 was reached. The fermented milk was heat treated at 75°C for 25 seconds. The samples were refrigerated in sterile cups / bottles before use. AI (Chr. Hansen A / S) + / - 0.1% fructose was aseptically added to each cup before equilibration to 25°C. The samples were thoroughly shaken (for bottles, use a sterile spoon or mixer) without introducing air. The second fermentation was at 25°C until the pH reached 4.3 or slightly lower. At the end of the second fermentation (day 1), the samples were stored at 25°C or 37°C and analyzed at a later time.

[0101] Milk Base 1 to Milk Base 4 were produced by adding sugar alcohols to the following composition (92.1% milk, 1.4% H2O, 0.9% sucrose, 1.5% modified starch and 0.1% pectin) according to the table below.

[0102] Table 7: Sample setup.

[0103]

[0104] Cell counts were measured over time as colony forming units (cfu) per gram (g) of sample stored at 25°C or 37°C.

[0105] Table 8a: Cell counts in samples stored at 25°C.

[0106]

[0107] *TNTC=Numerous

[0108] Table 8b: Cell counts in samples stored at 37°C.

[0109]

[0110] *TNTC=Numerous

[0111] The post-acidification of the samples was tracked over time. Adding fermentable sugars together with LGG could potentially lead to unwanted post-acidification. The effect of additional fructose depended on the milk base and, therefore, the presence of sugar alcohols. As can be seen in the table below, the post-acidification was lowest when fructose and LGG were added in MB3 (Ery+Xyl) and MB4 (Ery+Bal), measured at 25°C and 37°C.

[0112] Table 9a: pH in samples stored at 25°C.

[0113]

[0114] Table 9b: pH in samples stored at 37°C.

[0115]

[0116] The formation of CO2 in the sample was tracked over time. Briefly, the sample was placed in a glass bottle with a rubber stopper. The cap was pierced with a needle and the CO2 was measured using a portable headspace gas analyzer. CheckPoint3 measures the %CO2 in the bottle's headspace.

[0117] The results in the table below show that by adding fructose together with LGG, the production of CO2 can be delayed. This delay depends on the milk base and therefore on the presence of sugar alcohols.

[0118] Table 10a: % CO2 in samples stored at 25°C.

[0119]

[0120] Table 10b: % CO2 in samples stored at 37°C.

[0121]

[0122] The conversion of citric acid to acetic acid in the samples was followed over time.

[0123] The results in the table below show that by adding fructose together with LGG, the conversion of citric acid to acetic acid could be delayed at day 28. This delay depended on the storage temperature, the milk base and therefore on the presence of sugar alcohols.

[0124] Table 11a: Citric acid consumption in samples stored at 25°C.

[0125]

[0126] Table 11b: Acetic acid production in samples stored at 25°C.

[0127]

[0128] Table 11c: Citric acid consumption in samples stored at 37°C.

[0129]

[0130] Table 11d: Acetic acid production in samples stored at 37°C.

[0131]

[0132] Example 8 - Effect of pH on the Effect of Sugar Alcohols.

[0133] We investigated the effect of the pH of pre-acidified milk on the conversion of citric acid to acetic acid by strains of the genus Lactobacillus in the presence or absence of sweeteners. Milk pre-acidified to four different pH values (4.5; 5.0; 5.5; 6.0) was prepared by adding HCl to milk supplemented with 0.1% glucose (GLC) and + / - 2% xylitol (XYL). The pre-acidified milk was inoculated with either Lactobacillus rhamnosus (DVS LGG, Chr. Hansen) or Lactobacillus paracasei (DSM 16572). Each strain was grown in 2 mL of the different milks in 96-deep-well plates at 37°C for 3 days, after which the samples were stored at 30°C. After 3, 14, and 28 days, the milk was analyzed for citric acid and acetic acid content (g / L) by HPLC.

[0134] The results in the table below are consistent with those in Example 4, indicating that citric acid consumption in unacidified milk is faster than that in pre-acidified milk. Furthermore, the effect of the sweetener was observed after 28 days at pH 4.5, but only after 3 days at pH 5. No effect of the sweetener was observed above pH 5.0.

[0135] Table 12a: Effect of pH on citric and acetic acid concentrations (g / L) in LGG-inoculated milk + 0.1% GLC + / - 2% XYL.

[0136]

[0137] Table 12b: Effect of pH on citric and acetic acid concentrations (g / L) in milk inoculated with DSM 16573 + 0.1% GLC + / - XYL.

[0138]

[0139] The present invention has been described with reference to various embodiments, aspects, examples, etc. It is undesirable to understand these elements in isolation from each other. Therefore, the present disclosure provides a combination of two or more embodiments, aspects, examples, etc. All embodiments described herein are intended to fall within the scope of the present disclosure. By reference to the following detailed description of the entire specification, those skilled in the art will readily understand these and other embodiments of the present invention, and the present invention is not limited to any specific preferred embodiment disclosed.

Claims

1. A method for producing a fermented dairy product, comprising the following steps: a) adding one or more sweeteners to a dairy base or pre-acidified milk, wherein the dairy base is acidified until a first target pH of not more than 4.7 is reached, to obtain the pre-acidified milk; b) optionally heat treating the pre-acidified milk; c) inoculating the pre-acidified milk with lactose-deficient lactic acid bacteria until a second target pH of no more than 4.6 is reached to obtain the dairy product, wherein the first target pH is equal to or higher than the second target pH; and wherein conversion of citric acid to acetic acid in the dairy product during storage is delayed compared to a dairy product produced without the one or more sweeteners.

2. The method according to claim 1, wherein the lactose-deficient lactic acid bacteria in step (c) are strains of the genus Lactobacillus.

3. The method according to claim 2, wherein the strain of the genus Lactobacillus is selected from the group consisting of rhamnosus, casei and paracasei species.

4. The method according to any one of claims 1 to 3, wherein the lactose-deficient lactic acid bacteria in step (c) ferment the pre-acidified milk to obtain a fermented dairy product.

5. The method according to any one of claims 1 to 4, wherein the one or more sweeteners are sugars and / or sugar alcohols.

6. The method of claim 5, wherein the sugar is selected from the group consisting of fructose, galactose, glucose and sucrose.

7. The method of any one of claims 5-6, wherein the concentration of the sugar is 0.05%, 0.10%, 0.15%, 0.20%, 0.25%, 0.30%, 0.35%, 0.40%, 0.45% or 0.50% or within the range of 0.05%-0.50%, 0.10%-0.40% or 0.15%-0.30%.

8. The method of claim 5, wherein the one or more sweeteners are selected from C4 sugar alcohols [C4H 10 O4] and C5 sugar alcohols [C5H 12 O5] sugar alcohol.

9. The method of claim 8, wherein the C4 sugar alcohol is erythritol, D-threitol, or L-threitol; and the C5 sugar alcohol is xylitol, ribitol, D-arabitol, L-arabitol, D-lyxitol, or L-lyxitol.

10. The method of any one of claims 5, 8-9, wherein the concentration of the sugar alcohol is 0.5%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5% or 0.50% or is within the range of 0.5%-5.0%, 1.0%-4.5%, 1.5%-4.0%, 2.0%-3.5% or 2.5%-3.0%.

11. The method according to any one of the preceding claims, wherein the pre-acidification of the milk base in step (a) is chemical acidification or fermentation by one or more lactic acid bacteria.

12. The method according to claim 11, wherein the pre-acidification of the milk base in step (a) is due to fermentation by one or more lactic acid bacteria, and wherein the pre-acidified milk is heat-treated.

13. The method according to any one of claims 11-12, wherein the one or more lactic acid bacteria belong to the genus Streptococcus, such as S. thermophilus, or to the genus Lactobacillus, such as L. delbrueckii subsp. bulgaricus.

14. The method according to any one of claims 11 to 13, wherein the one or more lactic acid bacteria are lactose deficient.

15. The method according to claim 14, wherein the lactose-deficient strain is selected from the group consisting of: DSM 28952, DSM 28953, DSM 28910, DSM 32600 and DSM 32599.

16. An acidified or fermented dairy product obtainable by the method according to any one of claims 1 to 15.

17. The dairy product according to any one of claims 1 to 16, wherein the product is a dairy product, such as yogurt (set or stirred); Greek yogurt; yogurt-based products such as fruit yogurt, and yogurt-based beverages such as drinking yogurt; buttermilk; kefir; skimmed yogurt, quark yogurt.

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