Synergistic antimicrobial effects between rosemary extract, cultured glucose, and buffered vinegar

Abstract

The present invention relates to novel synergistic compositions comprising known "clean label" antimicrobial substances and methods for stabilizing food substances against typical foodborne spoilage microorganisms and / or pathogens. Another object of the present invention is to provide novel combinations of "clean label" antimicrobial agents that can be used in such methods.

Description

Field of the Invention

[0001] The present invention relates to antimicrobial methods and novel antimicrobial compositions that can be used in such methods. In order to find a new and powerful "clean label" antimicrobial solution, three different categories of antimicrobial agents, including plant extracts (such as plant essential oils), cultured glucose and buffered vinegar, were tested for their synergistic antimicrobial effects. When rosemary extract or rosemary essential oil, cultured glucose and buffered vinegar were combined, a surprising synergistic effect was observed. Combinations of two and / or three of these antimicrobial agents all showed synergistic antimicrobial effects against Salmonella entericia subsp. enteric serovar Typhimurium. Background of the Invention

[0003] Traditionally, chemical preservatives in combination with various processing aids are applied to food systems to prevent food spoilage microorganisms or foodborne pathogens. However, the growing consumer demand for natural and additive-free foods has prompted the food industry to seek effective "clean label" antimicrobial solutions to maintain food safety and stability.

[0004] Culture glucose is a commercially available food additive, which is fermented and prepared by sugar sources (such as corn, sucrose) or dairy-based sources (including skim milk). Culture glucose consists of various fermentation metabolites, wherein the main active ingredients are antimicrobial peptides and organic acids. The microorganisms involved in the fermentation are mainly probiotics, such as propionibacterium (including Propionibacterium freudenreichii) and lactic acid bacteria (including Lactococcus lactis). The main purpose of culture glucose is to inhibit the growth of bacteria, yeast and mold in food systems. In various applications including dairy products, baked goods or cooking products, culture glucose can be an effective preservative. However, when used in an effective dose, the product usually brings delicious characteristics to the product.

[0005] Growing evidence indicates that plant materials have the potential to exhibit antimicrobial activity, which has prompted researchers to investigate different extracts to inhibit bacteria, yeasts, and molds in various applications. Plant secondary metabolites are natural compounds known for their role in plant defense mechanisms and possess a variety of biological activities, including antimicrobial effects. They include alkaloids, flavonoids, tannins, terpenes, quinones, and resins. One of the major limitations of using plant extracts as antimicrobial agents in the food and feed industry is that plant extracts usually require high effective doses, which can negatively affect the sensory properties of the matrix to which they are applied.

[0006] Essential oils are mixtures of volatile compounds extracted from plant biomass and have been identified as natural antimicrobial agents, with their activity primarily thought to derive from phenolic components. However, the use of essential oils as food preservatives is limited by their high flavor profile, which can negatively affect the sensory properties of the food to which they are applied.

[0007] Buffered vinegar is another “clean label” alternative to chemical preservatives currently used in the food industry, especially for meat and poultry. Buffered vinegar is known to inhibit the growth of foodborne pathogens and common spoilage organisms. The taste impact of vinegar can be reduced by buffering the vinegar with a sodium or potassium base. However, the concentrations of buffered vinegar required to demonstrate antimicrobial activity often produce a “vinegar” or “acidic” character in the final product and negatively impact sensory properties.

[0008] The present invention identifies synergistic combinations that can effectively inhibit the growth of spoilage microorganisms or pathogens at lower concentrations, thereby reducing the associated flavor impact of the component antimicrobial agents. These synergistic combinations can be used in meat and poultry, sauces and dressings, salads, hummus, seafood, cosmetics and / or nutritional supplements.

[0009] Purpose of the Invention

[0010] It is an object of the present invention to provide novel synergistic compositions comprising known "clean label" antimicrobial substances, and methods for stabilizing food substances against typical foodborne spoilage microorganisms and / or pathogens. The synergistic compositions and methods of the present invention are intended to replace current synthetic antimicrobials and methods utilizing synthetic antimicrobials, such as sodium diacetate, potassium lactate, and sodium benzoate. Another object of the present invention is to provide novel combinations of "clean label" antimicrobials that can be used in such methods. SUMMARY OF THE INVENTION

[0012] The present invention relates to an antimicrobial composition comprising at least two antimicrobial ingredients selected from plant extracts (including plant essential oils), cultured glucose and buffered vinegar, wherein the composition exhibits synergistic antimicrobial activity.

[0013] Another aspect of the present invention relates to such an antimicrobial composition, wherein the plant extract component is a rosemary extract.

[0014] Another aspect of the present invention relates to such an antimicrobial composition, wherein the rosemary extract is a deodorized rosemary extract having a low essential oil content (eg, 0.06-0.63% w / w).

[0015] Another aspect of the present invention relates to such an antimicrobial composition, wherein the plant extract component is a plant essential oil.

[0016] Another aspect of the present invention relates to such an antimicrobial composition, wherein the plant essential oil component is rosemary essential oil.

[0017] Another aspect of the present invention relates to such antimicrobial compositions, wherein the culture glucose component is a fermentation product of a sugar source (such as corn, sucrose) or a dairy-based source (including skim milk).

[0018] Another aspect of the present invention relates to antimicrobial compositions wherein the buffered vinegar component is a fermentation product of corn and sucrose, but may also be adapted to conventional vinegar mixtures having acetic acid as a major component.

[0019] In one embodiment, the buffered vinegar component is selected from a common vinegar mixture having acetic acid as the main component.

[0020] In one embodiment, the antimicrobial composition comprises rosemary extract and cultured glucose.

[0021] In one embodiment, the antimicrobial composition comprises rosemary extract and buffered vinegar.

[0022] In one embodiment, the antimicrobial composition comprises rosemary essential oil and cultured glucose.

[0023] In one embodiment, the antimicrobial composition comprises rosemary essential oil and buffered vinegar.

[0024] In one embodiment, the antimicrobial composition comprises cultured glucose and a buffered ester.

[0025] In one embodiment, the antimicrobial composition comprises rosemary extract, cultured glucose, and buffered vinegar.

[0026] In one embodiment, the antimicrobial composition comprises rosemary essential oil, cultured dextrose, and buffered vinegar.

[0027] Another aspect of the present invention relates to a stabilized food, beverage, cosmetic and / or nutritional supplement comprising the antimicrobial composition.

[0028] Another aspect of the present invention relates to a method for stabilizing a food, beverage, cosmetic and / or nutritional supplement comprising incorporating an effective amount of an antimicrobial composition, wherein the composition exhibits synergistic antimicrobial activity.

[0029] In one embodiment of the method, the antimicrobial composition comprises two antimicrobial ingredients.

[0030] In one embodiment of the method, the antimicrobial composition comprises three antimicrobial ingredients. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] Figure 1 shows the antimicrobial testing of rosemary essential oil (RO) and cultured glucose (CD).

[0033] Figure 2 shows the antimicrobial testing of rosemary essential oil (RO) and buffered vinegar (BV).

[0034] FIG3 shows the antimicrobial testing of culture glucose (CD) and buffered vinegar (BV).

[0035] Figure 4 shows the antimicrobial testing of rosemary essential oil (RO), cultured glucose (CD) and buffered vinegar (BV).

[0036] 5 shows antimicrobial testing of A) rosemary oil (RO), cultured dextrose (CD), and a combination of RO and CD; B) rosemary oil (RO), buffered vinegar (BV), and a combination of RO and BV; and C) cultured dextrose (CD), buffered vinegar (BV), and a combination of CD and BV.

[0037] Figure 6 shows antimicrobial testing of A) buffered vinegar (BV) and bell pepper leaf oil (PO); B) cultured glucose (CD) and clove oil (CO); and C) cultured glucose (CD) and thyme oil (TO).

[0038] Figure 7 shows the synergistic antimicrobial activity of encapsulated rosemary essential oil, cultured glucose, and buffered vinegar against a Salmonella cocktail in fresh ground turkey.

[0039] FIG8 shows the synergistic antimicrobial activity of encapsulated rosemary essential oil, cultured glucose, and buffered vinegar against coliform bacteria in fresh ground turkey.

[0040] Figure 9 shows the synergistic antimicrobial activity of dried rosemary extract, cultured glucose, and buffered vinegar against total aerobic bacteria in fresh ground turkey.

[0041] Figure 10 shows the synergistic antimicrobial activity of dried rosemary extract, cultured glucose, and buffered vinegar against Listeria in chicken salad. DETAILED DESCRIPTION OF THE INVENTION

[0043] Microbial strains and culture conditions

[0044] Salmonella enterica serovar Typhimurium (ATCC 14028 TM) was obtained from ATCC (Manassas, VA). Alternatively, lactic acid bacteria including Propionibacterium, Lactococcus lactis, Lactobacillus reuteri can also be used as representative microbial sources for the test. Brain heart infusion broth (BD, Sparks, MD) was used as a growth medium and adjusted to pH 5.2 with HCl. Chloramphenicol (Sigma, St.Louis, MO) was used as a positive control. All cultures were incubated at 37°C for 24 hours under stirring at 220rpm. Growth was determined by measuring the optical density (OD600) at 600nm using a BIOTEK Power Wave HT 340 spectrophotometer (BIOTEK, Winooski, VT). Incremental growth was calculated by subtracting the growth value measured at 0 hour from the growth value measured at 24 hours.

[0045] Culture glucose is produced by fermentation of corn sugar or sucrose or an alternative sugar source.

[0046] Buffered vinegar is made by fermenting corn and cane sugar, but can also be used with regular vinegar mixtures that have acetic acid as the main ingredient.

[0047] Antimicrobial synergy determined

[0048] The percent inhibition was calculated by the following equation:

[0049]

[0050] All experiments were performed in triplicate.

[0051] Two-factor synergy test

[0052] Rosemary essential oil (RO), buffered vinegar (BV) and cultured glucose (CD) were tested for their synergistic antimicrobial activity against S. Typhimurium. Prior to synergy testing, the minimum inhibitory concentration (MIC) of each mixture was determined. Concentrations showing sub-inhibitory and non-inhibitory effects were used for synergy testing.

[0053] The two-factor synergy was determined by the following equation:

[0054] - Percent inhibition of [a% rosemary essential oil and b% cultured glucose mixture] > Percent inhibition of [a% rosemary essential oil] + Percent inhibition of [b% cultured glucose]

[0055] -Percent inhibition of [a% rosemary essential oil and c% buffered vinegar mixture] > Percent inhibition of [a% rosemary essential oil] + Percent inhibition of [c% buffered vinegar]

[0056] -[% inhibition of mixture of b% culture glucose and c% buffer vinegar] >% inhibition of [b% culture glucose] +% inhibition of [c% buffer vinegar]

[0057] Three-factor synergy test

[0058] The concentrations of rosemary essential oil, buffered vinegar, and cultured glucose used in the three-way synergy test were designed to have a slight synergistic antimicrobial effect when the two substances were combined, but did not show 100% inhibition against S. typhimurium.

[0059] when Meet all of the following conditions The synergy of three factors is determined:

[0060] - Percent inhibition of [a% rosemary oil and b% cultured glucose and c% buffered vinegar mixture] > Percent inhibition of [a% rosemary oil] + Percent inhibition of [b% cultured glucose] + Percent inhibition of [c% buffered vinegar]

[0061] -% inhibition of [a% rosemary oil and b% cultured glucose and c% buffered vinegar mixture] >% inhibition of [a% rosemary oil and b% cultured glucose mixture] +% inhibition of [c% buffered vinegar]

[0062] -% inhibition of [a% rosemary oil and b% cultured glucose and c% buffered vinegar mixture] >% inhibition of [a% rosemary oil and c% buffered vinegar mixture] +% inhibition of [b% cultured glucose]

[0063] -% inhibition of [a% rosemary essential oil and b% cultured glucose and c% buffered vinegar mixture] >% inhibition of [b% cultured glucose and c% buffered vinegar mixture] +% inhibition of [a% rosemary essential oil]

[0064] Antimicrobial Testing in Food Models

[0065] Microbial strains and culture conditions

[0066] Salmonella enterica subspecies Typhimurium (ATCC 14028 TM ), Salmonella enterica subspecies Typhimurium (ATCC 700720 TM ), Salmonella enterica subsp. enterica serovar Enteritidis (ATCC 4931 TM), Salmonella enterica subsp. enterica serovar Newport (ATCC 6962 TM ) was obtained from ATCC (Manassas, VA). The strains were initially grown in brain heart infusion broth (BD, Sparks, MD) at 37°C with agitation at 220 rpm for 24 hours before inoculation into the food matrix. All four strains of Salmonella were grown individually and then combined into a mixture before inoculation.

[0067] Listeria monocytogenes ATCC 19115 TM , Listeria monocytogenes ATCC 19115 TM , Listeria monocytogenes ATCC 19115 TM Obtained from ATCC (Manassas, VA). Strains were initially grown in Brain Heart Infusion Broth (BD, Sparks, MD) at 37°C with agitation at 220 rpm for 24 hours prior to inoculation into the food matrix. All three strains of Listeria monocytogenes were grown individually and then combined into a mixture prior to inoculation.

[0068] Ground Turkey Model Preparation (for Salmonella and Coliform Challenge Testing)

[0069] Fresh ground turkey meat (80% lean meat and 20% fat) was purchased from a local store and immediately transferred to the laboratory. The ground turkey meat was coarsely ground (16 mm) and then finely ground (5 mm). The antimicrobial mixture was directly added to the ground turkey meat in a sterile homogenizing bag and homogenized twice with a homogenizer (Seward stomacher 400Circulator) at 230 rpm, each for 1 minute.

[0070] Chicken salad model preparation (for Listeria challenge testing)

[0071] Fresh chicken breast, celery, mayonnaise, salt, black pepper were purchased from a local store and transferred to the laboratory. Chicken breast was boiled in hot water until the internal temperature reached 74°C and cut into 2 cm cubes. Celery was washed with cold water, dried, and cut into 1 cm pieces. A chicken salad model was prepared by mixing 1640 g of cooked chicken cubes, 105 g of celery, 180 g of mayonnaise, 2 g of black pepper, and 10 g of salt. The antimicrobial mixture was added directly to the chicken salad in a sterile homogenizing bag and homogenized twice at 230 rpm for 1 minute each time using a homogenizer (Seward stomacher 400Circulator).

[0072] Microbiological sampling

[0073] The food sample (25 grams) was transferred to a sterile homogenizing bag and filled to 250 grams with 0.1% peptone water. The mixture was homogenized at 230 rpm for 1 minute and serially diluted to the appropriate level, and then the dilution was plated on a selective agar medium, followed by incubation at 37°C for 24 hours. In order to count the genus Salmonella, the sample dilution was plated on an XLD (xylose lysine deoxycholic acid) agar medium, and the red colonies with a black center were counted. The yellow colonies that appear on XLD agar are Gram-negative bacteria and lysine decarboxylase-negative bacteria, and are considered to be coliforms. The total plate count was determined by plating the sample on a total plate count (TPC) agar and incubating at 30°C for 24 hours. The monocytogenes Listeria count was determined by plating the sample on Listeria selective agar (LSA) and incubating at 37°C for 24 to 48 hours. Example

[0074] The following examples illustrate the invention without limiting its scope.

[0075] Example 1 Two-factor synergy between rosemary essential oil and cultured glucose

[0076] Salmonella enterica serovar Typhimurium (ATCC 14028 TM ) Brain heart infusion broth was used as growth medium, cultured at 37°C under stirring at 220 rpm for 24 hours, and pH was adjusted to pH 5.2 with HCl. Chloramphenicol was used as a positive control. Growth was determined by measuring the optical density at 600 nm. Incremental growth was calculated by subtracting the growth measured at 0 hours from the growth measured at 24 hours. As shown below and in Figure 1, test antimicrobial agents and their percentage inhibition are provided:

[0077]

[0078] Example 2 Two-factor synergy between rosemary essential oil and buffered vinegar

[0079] Salmonella enterica serovar Typhimurium (ATCC 14028 TM ) Brain heart infusion broth was used as growth medium, cultured at 37°C under stirring at 220 rpm for 24 hours, and pH was adjusted to pH 5.2 with HCl. Chloramphenicol was used as a positive control. Growth was determined by measuring the optical density at 600 nm. Incremental growth was calculated by subtracting the growth measured at 0 hours from the growth measured at 24 hours. As shown below and in Figure 2, test antimicrobial agents and their percentage inhibition are provided:

[0080]

[0081] Example 3 Cultivation of two-factor synergy between glucose and buffered acetic acid

[0082] Salmonella enterica serovar Typhimurium (ATCC 14028 TM ) Brain heart infusion broth was used as growth medium, cultured at 37°C under stirring at 220 rpm for 24 hours, and pH was adjusted to pH 5.2 with HCl. Chloramphenicol was used as a positive control. Growth was determined by measuring the optical density at 600 nm. Incremental growth was calculated by subtracting the growth measured at 0 hours from the growth measured at 24 hours. As shown below and in Figure 3, test antimicrobial agents and their percentage inhibition are provided:

[0083]

[0084] Example 4 Three-factor synergy between rosemary essential oil, cultured glucose and buffered vinegar

[0085] To evaluate the three-factor synergy between the corresponding antimicrobial test substances, the concentrations of these substances were reduced and the three-factor synergy was compared with the two-factor synergy between the test antimicrobials using reduced concentrations. 14028 TM ) Brain heart infusion broth was used as growth medium, cultured at 37°C under stirring at 220 rpm for 24 hours, and pH was adjusted to pH 5.2 with HCl. Chloramphenicol was used as a positive control. Growth was determined by measuring the optical density at 600 nm. Incremental growth was calculated by subtracting the growth measured at 0 hours from the growth measured at 24 hours. As shown below and in Figures 4 and 5, test antimicrobial agents and their percentage inhibition are provided:

[0086]

[0087] Example 5 Two-factor synergy between various essential oils and combinations of cultured glucose and buffered vinegar

[0088] To evaluate the potential for synergy between the corresponding antimicrobial test substances and various other essential oils selected from clove, thyme and bell pepper leaf, sub-inhibitory concentrations of the additional essential oil extracts were evaluated alone and in combination with incubation glucose and buffered vinegar. Since the essential oils evaluated showed antagonism or non-synergistic effects on the inhibition observed with incubation glucose and buffered vinegar, the concentrations of incubation glucose and buffered vinegar were increased compared to the sub-inhibitory concentrations used in the three-factor synergy experiment with rosemary essential oil. 14028 TM ) Brain heart infusion broth was used as growth medium, cultured at 37°C under stirring at 220 rpm for 24 hours, and pH was adjusted to pH 5.2 with HCl. Chloramphenicol was used as a positive control. Growth was determined by measuring the optical density at 600 nm. Incremental growth was calculated by subtracting the growth measured at 0 hours from the growth measured at 24 hours. As shown below and in Figure 6, test antimicrobial agents and their percentage inhibition are provided:

[0089]

[0090] Example 6 Synergistic Antimicrobial Activity of Encapsulated Rosemary Essential Oil, Cultured Glucose and Buffered Vinegar Against Salmonella Mixture and Coliforms in Fresh Ground Turkey

[0091] Salmonella enterica subspecies Typhimurium (ATCC 14028 TM ), Salmonella enterica subspecies Typhimurium (ATCC 700720 TM ), Salmonella enterica subspecies enteritidis serotype (ATCC 4931 TM ), Salmonella enterica subspecies Newport serotype (ATCC 6962 TM) were cultured separately in brain heart infusion broth (BD, Sparks, MD) at 37°C with agitation at 220 rpm for 24 hours and then combined into a mixture. Fresh ground turkey was prepared and mixed with each antimicrobial treatment: 1) control treatment, no antimicrobial added (control), 2) 0.3% culture glucose and buffered vinegar mixture mixed in a 50:50 ratio (0.3% CDV), 3) 5% encapsulated rosemary essential oil, 20% loading yield (5% RO), and 4) 0.3% CDV + 5% RO. After adding the antimicrobial treatment, the Salmonella mixture was inoculated into the ground turkey, with a target initial population of approximately 4 log CFU / g. The ground turkey samples were then vacuum packed and stored at 10°C for a period of 14 days, and the Salmonella population was determined by plating the samples on XLD agar medium and counting red colonies with black centers. Yellow colonies appearing on XLD agar are Gram-negative and lysine decarboxylase-negative bacteria and are considered to be coliforms. All experiments were performed in duplicate and the average values ​​are reported.

[0092] Compared to the other treatments, ground turkey treated with 0.3% CDV + 5% RO showed the best results by very effectively inhibiting the growth of Salmonella. The results are shown in Figure 7. When comparing Day 14 (3.1 log CFU / g) and Day 0 (3.8 log CFU / g), a bactericidal effect (reduction in Salmonella counts) was observed. In particular, on Day 4, rosemary essential oil (5% RO) alone showed no inhibitory effect on Salmonella, but when rosemary essential oil was combined with cultured glucose and buffered vinegar, a clear synergistic antimicrobial effect was observed.

[0093] The combination of cultured glucose, buffered vinegar and encapsulated rosemary essential oil also performed best and showed a synergistic antimicrobial effect against coliforms. The results are shown in FIG8 .

[0094] Example 7 Synergistic Antimicrobial Activity of Dried Rosemary Extract, Cultured Glucose and Buffered Vinegar Against Spoilage Microorganisms in Fresh Ground Turkey

[0095] Fresh ground turkey (80% lean and 20% fat) was coarsely ground (16 mm) and then finely ground (5 mm) and mixed with each set of antimicrobial treatments: 1) a control treatment with no antimicrobial added (control), 2) a 1% cultured glucose and buffered vinegar mixture (CDV) mixed in a 50:50 ratio, 3) a 1% mixture of 2.2% dried rosemary extract, 85% cultured glucose, 13.85% maltodextrin and 0.7% silicon dioxide (NCD) and 4) a 1% mixture of 2.2% dried rosemary extract, 50% cultured glucose, 35% buffered vinegar, 13.85% maltodextrin and 0.7% silicon dioxide (1% NCDV). The ground turkey samples were vacuum packed and stored at 4°C for a period of 17 days and the total plate count was checked by plating the samples on total plate count (TPC) agar. All experiments were performed in duplicate and the average values ​​are reported.

[0096] 1% NCD was the most effective antimicrobial agent for inhibiting total aerobic bacteria. Although the mean of total plate counts for the 1% NCD treatment at day 5 showed a slight increase compared to day 0, the difference was not statistically significant. Ground turkey treated with the combination of NCD was more effective in inhibiting total aerobic bacterial growth compared to CDV without rosemary extract (Figure 9), showing a 1.6 log difference at day 17. On the other hand, the NCD treatment did not perform well compared to the other two treatments.

[0097] When dry rosemary extract was used alone at the same concentration (2.2%), no antimicrobial activity was observed (data not shown), demonstrating that the combination effect was synergistic.

[0098] To determine which microorganisms were inhibited by the additional synergistic effect of adding dried rosemary extract to the synergistic combination, colonies grown on TPC agar from ground turkey samples treated with 1% CDV but not from samples treated with 1% NCDV were isolated and all isolated colonies were identified as Pseudomonas by 16s rRNA sequencing.

[0099] Example 8 Synergistic Antimicrobial Activity of Dried Rosemary Extract, Cultured Glucose and Buffered Vinegar Against Listeria Monocytogenes in Chicken Salad

[0100] Listeria monocytogenes ATCC 19115 TM , Listeria monocytogenes ATCC 19115 TM , Listeria monocytogenes ATCC 19115 TMObtained from ATCC (Manassas, VA). Strains were initially grown in brain heart infusion broth (BD, Sparks, MD) at 37°C with agitation at 220 rpm for 24 hours and then inoculated into food matrices. All three strains of Listeria monocytogenes were grown individually and then combined into a mixture before inoculation. Chicken salad was prepared and mixed with each antimicrobial treatment: 1) a control treatment with no antimicrobial added (Control), 2) a 1% culture dextrose and buffered vinegar mixture mixed in a 50:50 ratio (CDV), 3) a 1% mixture of 2.2% dried rosemary extract, 85% culture dextrose, 13.85% maltodextrin, and 0.7% silicon dioxide (NCD), and 4) a 1% mixture of 2.2% dried rosemary extract, 50% culture dextrose, 35% buffered vinegar, 13.85% maltodextrin, and 0.7% silicon dioxide (1% NCDV). After adding the antimicrobial treatment, the Listeria cocktail was inoculated into the chicken salad, with a target initial population of approximately 4 log CFU / g. The chicken salad samples were stored at 4°C for a period of 28 days and the Listeria monocytogenes population was examined by plating the samples on LSA agar medium and counting black colonies. All experiments were performed in duplicate and the average values ​​are reported.

[0101] The combination of dried rosemary extract, cultured glucose, and buffered vinegar (NCDV) treatments performed better than the combination of cultured glucose and buffered vinegar (CDV) or the combination of dried rosemary extract and cultured glucose (NCD). At day 21, 1% NCD had higher L. monocytogenes counts compared to the other two treatments, and at day 28, 1% CDV and 1% NCD treatments had higher L. monocytogenes counts than 1% NCDV. When dried rosemary extract was used alone at the same concentration (2.2%), no antimicrobial activity was observed (data not shown), demonstrating that the combination effect was synergistic.

[0102] *****

[0103] The present invention is not limited to the scope of the specific embodiments described herein. In fact, various modifications of the present invention other than those described herein will become apparent to those skilled in the art from the foregoing description. These modifications are intended to fall within the scope of the appended claims.

[0104] All patents, applications, publications, test methods, literature, and other materials cited herein are incorporated by reference.

[0105] *****

Claims

1. An antimicrobial composition comprising rosemary essential oil, cultured glucose and buffered vinegar, wherein the rosemary essential oil is present at a concentration of 0.5% or less, the cultured glucose is present at a concentration of 0.112% or less, and the buffered vinegar is present at a concentration of 0.044% or less, wherein the composition exhibits synergistic antimicrobial activity.

2. The antimicrobial composition of claim 1, wherein the culture glucose component is a sugar source or a fermentation product of a dairy-based source.

3. The antimicrobial composition of claim 2, wherein the sugar source comprises corn and sucrose.

4. The antimicrobial composition of claim 2, wherein the dairy-based source comprises skim milk.

5. The antimicrobial composition according to any one of claims 1 to 4, wherein the buffered vinegar component is a fermentation product of corn and sucrose, or a common vinegar mixture having acetic acid as a main component.

6. An antimicrobial composition according to any one of claims 1 to 4, wherein the buffered vinegar component is selected from common vinegar mixtures having acetic acid as a major component.

7. A stabilized food, beverage, cosmetic and / or nutritional supplement comprising the antimicrobial composition according to any one of claims 1 to 6.

8. A method for stabilizing food, beverages, cosmetics and / or nutritional supplements comprising incorporating an effective amount of an antimicrobial composition according to any one of claims 1 to 6, wherein the composition exhibits synergistic antimicrobial activity.

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