Bacterial, fungal, and yeast growth inhibitor containing allulose

Abstract

Disclosed herein are a microorganism growth inhibitor including allulose-containing saccharides and a fermented alcoholic beverage comprising the same.

Claims

1. A fermented alcoholic beverage comprising: (a) at least one microorganism selected from the group consisting of Lactobacillus casei and Lactococcus lactis subsp. lactis; and (b) a growth inhibitor for a microorganism, comprising: allulose-containing saccharides; wherein the microorganism comprises at least one selected from the group consisting of Lactobacillus casei and Lactococcus lactis subsp. lactis; wherein the allulose is present in an amount of 50 parts by weight to 100 parts by weight relative to 100 parts by weight of the saccharides based on dried solid content.

2. The fermented alcoholic beverage according to claim 1, wherein the fermented alcoholic beverage is makgeolli or wine.

3. The fermented alcoholic beverage according to claim 1, wherein the fermented alcoholic beverage is fermented using at least one microorganism selected from the group consisting of Lactobacillus casei and Lactococcus lactis subsp. lactis.

4. The fermented alcoholic beverage of claim 1, wherein the saccharides are free from sucrose, glucose, or a combination thereof.

5. The fermented alcoholic beverage according to claim 4, wherein the fermented alcoholic beverage is makgeolli or wine.

6. The fermented alcoholic beverage according to claim 4, wherein the fermented alcoholic beverage is fermented using at least one microorganism selected from the group consisting of Lactobacillus casei and Lactococcus lactis subsp. lactis.

Description

BRIEF DESCRIPTION OF DRAWINGS

(1) FIGS. 1 and 2 are graphs showing time-dependent CFU values for bacterial targets (Lactobacillus casei (FIG. 1) and Lactococcus lactis subsp. lactis (FIG. 2)), as measured on samples of Comparative Examples using glucose or sucrose as saccharides and samples of Examples using allulose as saccharides.

(2) FIGS. 3 to 7 are graphs showing time-dependent CFU values for fungus targets (Aspergillus oryzae (FIG. 3), Aspergillus awamori (FIG. 4), Monascus purpureus (FIG. 5), Monascus ruber (FIG. 6), and Rhizopus oryzae (FIG. 7)), as measured on samples of Comparative Examples using glucose or sucrose as saccharides and samples of Examples using allulose as saccharides.

(3) FIGS. 8 and 9 are graphs showing time-dependent CFU values for yeast targets (Saccharomyces cerevisiae (FIG. 8) and Saccharomyces pastorianus (FIG. 9)), as measured on samples of Comparative Examples using glucose or sucrose as saccharides and samples of Examples using allulose as saccharides.

DETAILED DESCRIPTION OF THE INVENTION

(4) Hereinafter, the present invention will be described in more detail with reference to examples. However, it should be noted that these examples are provided for illustration only and should not be construed in any way as limiting the invention. In addition, these examples are provided for more complete understanding of the present invention by one of ordinary skill in the art.

EXAMPLES

Experimental Example 1: Preparation of Microorganism and Medium for Fermentation of Fermented Alcoholic Beverage

(5) In order to evaluate inhibition of post-fermentation of fermented alcoholic beverages, a degree of inhibition of the growth of microorganisms commonly used in fermented alcoholic beverages was determined.

(6) Specifically, each of the microorganisms (bacteria, fungi, and yeasts) obtained from Korean Culture Center of Microorganisms (KCCM) was subcultured three or four times in a culture medium (Table 1) by a typical method to enhance activity. Then, the cultured microorganism was inoculated into a culture medium (Table 1), followed by inducing over-cultivation to a concentration of 10.sup.5 CFU/mL or more, thereby obtaining a strain mother liquor (Table 1).

(7) TABLE-US-00001 Table 1 Item Name KCCM ATCC Culture medium Bacterium Lactobacillus casei 12452 393 LACTOBACILI MRS (MRS) Lactococcus lactis 41573 15577 LACTOBACILI subsp. lactis MRS (MRS) Fungus Aspergillus oryzae 12698 1011 POTATO DEXTROSE BROTH (PDB) Aspergillus 60247 14331 POTATO awamori DEXTROSE BROTH (PDB) Monascus 35473 16365 POTATO purpureus DEXTROSE BROTH (PDB) Monascus ruber 11845 16378 POTATO DEXTROSE BROTH (PDB) Rhizopus oryzae 11697 4858 POTATO DEXTROSE BROTH (PDB) Yeast Saccharomyces 11304 9080 YEAST MOLD cerevisiae BROTH (YMB) Saccharomyces 11523 2345 YEAST MOLD pastorianus BROTH (YMB)

Experimental Example 2: Confirmation for Growth of Microorganism Depending on Composition of Saccharides

(8) In order to compare degrees of inhibition of the growth of microorganisms depending upon the kind of purely added saccharides without a bias due to alcohol and other additives contained in a fermented alcoholic beverage, only the kind of saccharides added to a culture medium for each of the microorganisms was varied and investigated.

(9) Specifically, as saccharides of a culture medium suited to each strain, water-containing crystalline glucose (purity 98 wt % or more, CJ Cheiljedang), sucrose (purity 98 wt % or more, CJ Cheiljedang), and crystalline allulose (purity: 98 wt % or more, CJ Cheiljedang) were added to each of the culture medium, thereby preparing a modified culture medium (Tables 2 to 4). Then, 100 mL of the prepared modified culture medium was inoculated with 1 mL of the mother liquor of each strain prepared in Experimental Example 1, followed by observation of time-dependent changes in cell number of each strain. That is, MRS media as listed in Table 2, PDB media as listed in Table 3, and YMB media as listed in Table 4 were inoculated with bacteria (Lactobacillus casei and Lactococcus lactis subsp. lactis), fungi (Aspergillus oryzae, Aspergillus awamori, Monascus purpureus, Monascus ruber, and Rhizopus oryzae), and yeasts (Saccharomyces cerevisiae and Saccharomyces pastorianus), respectively, in the same manner. Then, each of the media inoculated with the bacteria was stored at 37° C. for 12, 24 and 48 hours to be used as a specimen for measurement of the number of single cell colonies, and each of the media inoculated with the fungi or yeasts was stored at 25° C. for 24, 48, and 72 hours to be used as a specimen for measurement of the number of single cell colonies.

(10) Measurement of the number of single cell colonies was carried out by a typical microorganism test. Specifically, 1 mL of each specimen was gradually diluted with 9 mL of 0.9% sterilized physiological saline by a 10-fold dilution method, thereby obtaining a diluted specimen. Thereafter, 1 mL of the diluted specimen was placed in a petri dish and 25 mL of a normal culture medium with agar added thereto was poured into the petri dish so as to be sufficiently mixed and solidified, thereby obtaining an inoculated specimen. Here, as the normal culture medium with agar added thereto, MRSA, PDA, and YMA were used for the bacteria, the fungi, and the yeasts, respectively.

(11) Thereafter, the measured number of cell colonies was multiplied by a dilution factor, thereby calculating a value of colony forming unit (CFU) per mL of sample.

(12) TABLE-US-00002 TABLE 2 Bacterium Comparative Comparative Example 1-1 Example 1-2 Example 1 Protease peptone, Protease peptone, Protease peptone, 10.0 g 10.0 g 10.0 g Beef extract, 8.0 g Beef extract, 8.0 g Beef extract, 8.0 g Yeast extract, 4.0 g Yeast extract, 4.0 g Yeast extract, 4.0 g Polysorbate Polysorbate Polysorbate (Tween 80), 1.0 g (Tween 80), 1.0 g (Tween 80), 1.0 g Ammonium Ammonium Ammonium citrate, 2.0 g citrate, 2.0 g citrate, 2.0 g Sodium acetate, 5.0 g Sodium acetate, 5.0 g Sodium acetate, 5.0 g K.sub.2HPO.sub.4, 2.0 g K.sub.2HPO.sub.4, 2.0 g K.sub.2HPO.sub.4, 2.0 g MgSO.sub.4 * 7H.sub.2O, 0.1 g MgSO.sub.4 * 7H.sub.2O, 0.1 g MgSO.sub.4 * 7H.sub.2O, 0.1 g MnSO.sub.4 * 4H.sub.2O, 0.05 g MnSO.sub.4 * 4H.sub.2O, 0.05 g MnSO.sub.4 * 4H.sub.2O, 0.05 g Distilled water, Distilled water, Distilled water, balance of 1.0 L balance of 1.0 L balance of 1.0 L Glucose, 20.0 g Sucrose, 20.0 g Allulose, 20.0 g

(13) TABLE-US-00003 TABLE 3 Fungus Comparative Comparative Example 2-1 Example 2-2 Example 2 POTATO POTATO POTATO STARCH, 4.0 g STARCH, 4.0 g STARCH, 4.0 g Distilled water, Distilled water, Distilled water, balance of 1.0 L balance of 1.0 L balance of 1.0 L Glucose, 20.0 g Sucrose, 20.0 g Allulose, 20.0 g

(14) TABLE-US-00004 TABLE 4 Yeast Comparative Comparative Example 3-1 Example 3-2 Example 3 Yeast extract, 3.0 g Yeast extract, 3.0 g Yeast extract, 3.0 g Malt extract, 3.0 g Malt extract, 3.0 g Malt extract, 3.0 g Peptone, 5.0 g Peptone, 5.0 g Peptone, 5.0 g Distilled water, Distilled water, Distilled water, balance of 1.0 L balance of 1.0 L balance of 1.0 L Glucose, 10.0 g Sucrose, 10.0 g Allulose, 10.0 g

(15) As a result, it was confirmed that the number of cell colonies of the nine strains of Examples 1 to 3 using allulose as saccharides was significantly lower than that of the strains of Comparative Examples. In addition, it was confirmed that the strains of Comparative Examples 1-1, 2-1, and 3-1 using glucose as saccharides as in a typical culture medium exhibited a high degree of growth of microorganisms and the strains of Comparative Examples 1-2, 2-2, and 3-2 using sucrose as saccharides exhibited a degree of growth of microorganisms not significantly different from that of the strains inoculated using glucose as saccharides.

(16) Specifically, Lactobacillus casei and Lactococcus lactis subsp. lactis, as bacteria, in a culture medium using glucose as saccharides (Comparative Example 1-1) and a culture medium using sucrose as saccharides (Comparative Example 1-2) were increased in cell number by 40.6 to 137 times. Conversely, the bacteria in a culture medium using allulose as saccharides were increased in cell number only by 1.3 to 2.2 times. Therefore, substantially no strain growth was observed (FIGS. 1 and 2).

(17) In addition, Aspergillus oryzae, Aspergillus awamori, Monascus purpureus, Monascus ruber and Rhizopus oryzae, as fungi, were increased in cell number by 9.8 to 148.5 times in a culture medium using glucose as saccharides (Comparative Example 2-1) and 5.9 to 81.1 times in a culture medium using sucrose as saccharides (Comparative Example 2-2), respectively. Conversely, the fungi were increased in cell number only by 1.6 to 9.8 times in a culture medium using allulose as saccharides (Example 2). Therefore, statistically significant inhibition of microorganism growth was observed (FIGS. 3 to 7).

(18) Finally, Saccharomyces cerevisiae and Saccharomyces pastorianus, as yeasts, in a culture medium using glucose as saccharides (Comparative Example 3-1) and a culture medium using sucrose as saccharides (Comparative Example 3-2) were increased in cell number by 27.4 to 134.8 times. Conversely, the yeasts in a culture medium using allulose as saccharides were increased in cell number only by 1.3 to 3 times. Therefore, substantially no strain growth was observed (FIGS. 8 and 9).

(19) For each of the strains, the number of single colonies was measured after cultivation for 0, 12, 24, 48, and 72 hours in each of the culture media. Results are shown in Table 5.

(20) TABLE-US-00005 TABLE 5 Medium-dependent Culture medium Strain classification 0 hours 12 hours 24 hours 48 hours (saccharides) Lactobacillus Comparative 1.5 × 10.sup.7 1.7 × 10.sup.8 7.3 × 10.sup.8 1.1 × 10.sup.9 MRS casei Example 1-1 (glucose) Comparative 1.5 × 10.sup.7 4.5 × 10.sup.7 3.9 × 10.sup.8 4.0 × 10.sup.8 MRS Example 1-2 (sucrose) Example 1 1.5 × 10.sup.7 1.5 × 10.sup.7 1.8 × 10.sup.7 2.0 × 10.sup.7 MRS (allulose) Lactococcus Comparative 2.4 × 10.sup.7 7.0 × 10.sup.8 3.6 × 10.sup.9 3.3 × 10.sup.9 MRS lactis subsp. Example 1-1 (glucose) lactis Comparative 2.4 × 10.sup.7 1.8 × 10.sup.8 4.8 × 10.sup.8 9.7 × 10.sup.8 MRS Example 1-2 (sucrose) Example 1 2.4 × 10.sup.7 4.2 × 10.sup.7 4.6 × 10.sup.7 5.2 × 10.sup.7 MRS (allulose) Medium-dependent culture Strain classification 0 hours 24 hours 48 hours 72 hours medium Aspergillus Comparative 5.3 × 10.sup.8 8.7 × 10.sup.8 3.0 × 10.sup.9 5.2 × 10.sup.9 PDB oryzae Example 2-1 (glucose) Comparative 5.3 × 10.sup.8 1.0 × 10.sup.9 2.7 × 10.sup.9 3.1 × 10.sup.9 PDB Example 2-2 (sucrose) Example 2 5.3 × 10.sup.8 5.7 × 10.sup.8 1.9 × 10.sup.9 1.7 × 10.sup.9 PDB (allulose) Aspergillus Comparative 2.6 × 10.sup.7 7.2 × 10.sup.7 2.4 × 10.sup.9 3.3 × 10.sup.9 PDB awamori Example 2-1 (glucose) Comparative 2.6 × 10.sup.7 4.0 × 10.sup.7 1.3 × 10.sup.9 2.1 × 10.sup.9 PDB Example 2-2 (sucrose) Example 2 2.6 × 10.sup.7 4.8 × 10.sup.7 4.9 × 10.sup.8 8.6 × 10.sup.8 PDB (allulose) Monascus Comparative 6.4 × 10.sup.7 1.1 × 10.sup.8 9.9 × 10.sup.8 8.6 × 10.sup.8 PDB purpureus Example 2-1 (glucose) Comparative 6.4 × 10.sup.7 5.9 × 10.sup.7 3.8 × 10.sup.8 5.6 × 10.sup.8 PDB Example 2-2 (sucrose) Example 2 6.4 × 10.sup.7 6.3 × 10.sup.7 2.5 × 10.sup.8 3.1 × 10.sup.8 PDB (allulose) Monascus ruber Comparative 1.0 × 10.sup.7 6.8 × 10.sup.8 1.5 × 10.sup.9 8.0 × 10.sup.8 PDB Example 2-1 (glucose) Comparative 1.0 × 10.sup.7 7.7 × 10.sup.7 1.6 × 10.sup.8 2.4 × 10.sup.8 PDB Example 2-2 (sucrose) Example 2 1.0 × 10.sup.7 2.3 × 10.sup.7 1.0 × 10.sup.7 1.6 × 10.sup.7 PDB (allulose) Rhizopus oryzae Comparative 3.7 × 10.sup.6 1.1 × 10.sup.7 2.1 × 10.sup.8 1.3 × 10.sup.8 PDB Example 2-1 (glucose) Comparative 3.7 × 10.sup.6 7.4 × 10.sup.6 8.9 × 10.sup.7 1.2 × 10.sup.8 PDB Example 2-2 (sucrose) Example 2 3.7 × 10.sup.6 6.4 × 10.sup.6 2.5 × 10.sup.7 3.6 × 10.sup.7 PDB (allulose) Saccharomyces Comparative 3.5 × 10.sup.9 7.7 × 10.sup.9 .sup. 5.3 × 10.sup.10 .sup. 1.3 × 10.sup.11 YMB cerevisiae Example 3-1 (glucose) Comparative 3.5 × 10.sup.9 5.6 × 10.sup.9 .sup. 4.8 × 10.sup.10 .sup. 9.7 × 10.sup.10 YMB Example 3-2 (sucrose) Example 3 3.5 × 10.sup.9 3.9 × 10.sup.9 2.9 × 10.sup.9 4.5 × 10.sup.9 YMB (allulose) Saccharomyces Comparative 2.7 × 108 6.4 × 10.sup.9 .sup. 1.0 × 10.sup.10 .sup. 3.6 × 10.sup.10 YMB pastorianus Example 3-1 (glucose) Comparative 2.7 × 108 2.6 × 10.sup.9 2.6 × 10.sup.9 8.6 × 10.sup.9 YMB Example 3-2 (sucrose) Example 3 2.7 × 108 4.0 × 10.sup.8 8.9 × 10.sup.8 8.0 × 10.sup.8 YMB (allulose)

(21) In conclusion, it can be seen that, when allulose is used instead of sucrose commonly used as a sweetener added to fermented alcoholic beverages such as makgeolli, it is possible to reduce post-fermentation caused by growth of microorganisms that can occur during production or distribution of the fermented alcoholic beverages.

(22) Although some exemplary embodiments have been described herein, it should be understood by those skilled in the art that these embodiments are given by way of illustration only, and that various modifications, variations and alterations can be made without departing from the spirit and scope of the invention. Therefore, the embodiments and the accompanying drawings should not be construed as limiting the technical spirit of the present invention, but should be construed as illustrating the technical spirit of the present invention. The scope of the invention should be interpreted according to the following appended claims as covering all modifications or variations derived from the appended claims and equivalents thereof.