JP3364491B2 - Yogurt and method for producing the same - Google Patents

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a method for producing yogurt using yogurt and lactic acid bacteria.

[0002]

2. Description of the Related Art Yogurt is produced by adding lactic acid bacteria or yeast to milk and fermenting it, and its consumption as a health food has been increasing in recent years.

[0003] Yogurt is produced by different manufacturing methods.
Generally, it is roughly classified into a stationary yogurt and a stirring yogurt. The stationary yogurt is a coagulated yogurt, which is obtained by adding lactic acid bacteria to raw material milk, filling it in a container, and then fermenting it. The stationary yogurt includes plain yogurt containing only raw milk and no additives, and hard yogurt hardened into pudding with gelatin or agar with added sugar or flavor. On the other hand, the agitated yogurt is a yogurt in a fluid or liquid state, and the yogurt solidified in the tank is agitated into a fluid and then filled in a container. Stirred yogurt includes soft yogurt containing pulp and drink yogurt crushed into a liquid state.

It is known that when these yogurts are produced, the use of a suitable lactic acid bacterium in combination requires a shorter milk coagulation time than the use of only a single lactic acid bacterium. This utilizes the symbiotic phenomenon of lactic acid bacteria. Therefore, conventionally, when yogurt is produced by combining a plurality of lactic acid bacteria, the combination of lactic acid bacteria to be used has been selected such that the bacteria have a symbiotic relationship with each other, that is, a relationship that utilizes each other. For example, Lactobacillus bulgarix
icus) and Streptococcus thermophilus (S
treptococcus thermophilu
s) is the most frequently used combination of lactic acid bacteria,
The reason is that they are symbiotic with each other. The symbiotic relationship of lactic acid bacteria will be specifically described below by taking the case of producing yogurt using these as an example.

When Lactobacillus bulgaricus and Streptococcus thermophilus are added to milk, significant growth of Streptococcus thermophilus is observed in the early stage of fermentation. However, when PH decreases to about 5.5 to 5.0 with the increase in acidity due to the production of lactic acid, the growth of Streptococcus thermophilus becomes slow. On the other hand, the growth of Lactobacillus bulgaricus is promoted by the presence of carbon dioxide produced by Streptococcus thermophilus from formic acid and urea contained in milk under low oxygen. Then, Lactobacillus bulgaricus decomposes proteins during the fermentation process to produce many peptides and free amino acids. Among these free amino acids, valine, histidine, methionine, glutamic acid, and leucine act as growth-promoting substances of Streptococcus thermophilus whose activity is weakened, whereby the fermentative power of Streptococcus thermophilus is restored. As described above, the symbiotic relationship is a relationship in which lactic acid bacteria create factors that help each other and enhance mutual fermentation.

[0006] Further, the genus Bifidoba
Cerium) and Lactobacillus acidophilus (Lactobacillus acidop)
hilus) is often used as a mixture because of the same growth habit of the same enterobacteria. However, even in these cases, it has hitherto been an essential condition to select bacteria that have a relationship of helping each other or a relationship of not inhibiting the growth of each other.

As described above, in the conventional production of yogurt, it is common knowledge that the lactic acid bacteria to be used have a symbiotic relationship with each other, and there are some examples of producing yogurt using lactic acid bacteria having no symbiotic relationship. There wasn't.

[0008] An object of the present invention is to provide a yogurt by a combination of novel lactic acid bacteria, which is not bound by conventional common sense, and a method for producing the same.

Another object of the present invention is to provide a yogurt having excellent flavor and an intestinal regulating action, and a method for producing the same.

Other objects and advantages of the invention will be apparent from the description below.

[0011]

The present invention is directed to Lactococcus lactis subspecies lactis (La).
ctococcus lactis subsp. l
Lactococcus lactis subspecies cremoris (Lactococcus la)
ctis subsp. cremoris) and a bifidobacteria (Bifidobacterium) -containing yogurt.

Bifidobacte
The rium) can be B. animalis.

The present invention also provides a method for producing yogurt using lactic acid bacteria, wherein the lactic acid bacteria are Lactococcus lactis subspecies lactis (Lac
tococcus lactis subsp. la
ctis), Lactococcus lactis subspecies cremoris (Lactococcus lac)
tis subsp. cremoris) and a bifidobacteria genus (mixed bacterium).

The genus Bifidobacterium is Bifidum (B. bi
fidum), longum (B. longum), infantis (B. infantis), breve (B.
breve) and animalis (B. animali)
It can be at least one selected from the group consisting of s).

[0015]

BEST MODE FOR CARRYING OUT THE INVENTION Hereinafter, embodiments of the present invention will be described in detail with reference to the drawings.

The yogurt according to the present invention is characterized by containing Lactococcus lactis subspecies lactis, Lactococcus lactis subspecies cremoris and Bifidobacterium genus.

Further, the method for producing yogurt according to the present invention is a method for producing yogurt in which a lactic acid bacterium is added to ferment a protein, wherein the lactic acid bacterium is Lactococcus lactis subspecies lactis, Lactococcus lactis Sub Species
It is characterized by being a mixed bacterium consisting of Cremoris and Bifidobacteria.

FIG. 1 is a diagram showing an example of a process for producing yogurt according to the present invention. In the figure, the steps of mixing, homogenizing, sterilizing and cooling until lactic acid fermentation can be carried out according to conventional methods.

First, a protein as a raw material for yogurt is prepared. As the protein, in addition to milk protein derived from milk, goat milk, sheep milk, processed milk, skim milk, skim milk powder, whole milk powder, caseins and whey protein, etc., it is also possible to use vegetable protein and the like. it can. When skim milk powder or the like is used as the protein, water can be added as water. In addition, sweeteners such as sugar, fresh cream, fruits, fruit juice, agar, gelatin, oils and fats, etc.
A fragrance, a coloring agent or a stabilizer may be added to the raw material.

The raw materials and auxiliary raw materials to be used are weighed in predetermined amounts and then mixed. At this time, the mixture may be heated. For example, it can be heated at 80 ° C. to 85 ° C. for 15 minutes to 20 minutes. Next, sterilization treatment is performed after performing homogenization treatment. Sterilization treatment is, for example, 70 ° C to 90 ° C.
It can be performed by heating and holding for 5 to 30 minutes. Then, it cools to the fermentation temperature mentioned later. The mixture that has undergone the steps of mixing, homogenizing, sterilizing and cooling is hereinafter referred to as a mix.

In the present invention, Lactococcus lactis subspecies
It is characterized in that a mixed bacterium comprising lactis, Lactococcus lactis subspecies cremoris and Bifidobacterium is used.

Since Lactococcus lactis subspecies lactis and Lactococcus lactis subspecies cremoris have low biogenic ability, they are conventionally unsuitable for the production of yogurt which requires a rapid increase in acidity. It is a lactic acid bacterium that was considered appropriate.

Further, Lactococcus lactis subspecies lactis and Lactococcus lactis subspecies cremoris, and Bifidobacterium genus commonly used in the production of yogurt are
It is known that they do not have a symbiotic relationship with each other. That is,
It has been conventionally said that bifidobacteria hardly ferment when these are mixed and used for fermentation. The reason is considered as follows.

Bifidobacteria can grow under strong acidity, but due to the obligate anaerobic property, their growth is inhibited in an environment with a high oxygen content such as milk. Therefore, in general, it can be said that the addition of a growth promoting substance such as an oligosaccharide is indispensable for carrying out fermentation using the genus Bifidobacterium. On the other hand, Lactococcus lactis subspecies lactis and Lactococcus lactis subspecies cremoris can grow under oxygen, but have the property of being weak in strong acidity.

Thus, the genus Bifidobacterium and Lactococcus lactis subspecies lactis and Lactococcus lactis subspecies
The development environment that is compatible with Cremoris is different. Therefore, when these were mixed, it was thought that there was no clue of fermentation of the genus Bifidobacterium among lactic acid bacteria having no growth promoting substance and having different growth environments, and the productivity thereof was further reduced.

As described above, Lactococcus lactis subspecies lactis and Lactococcus lactis subspecies cremoris have no symbiotic relationship with the genus Bifidobacterium and are used in the conventional method for producing yogurt. It is a combination that was not made. However, as a result of diligent research, the present inventor has found that the yogurt according to the present invention can be obtained by mixing and using these bacteria.

Further, the present inventor has found that the yogurt according to the present invention can be obtained by setting the fermentation temperature in the optimum temperature range for the genus Bifidobacterium. Optimum temperature range for Bifidobacteria (30 ℃ -40 ℃)
Lactococcus lactis subspecies
Optimal temperature range for lactis and Lactococcus lactis subspecies cremoris (20 ° C
-30 ° C).

As described above, the yogurt of the present invention is suitable for the growth conditions which are generally preferable not only for the genus Bifidobacterium but also for Lactococcus lactis subspecies lactis and Lactococcus lactis subspecies cremoris. Is manufactured without. However, the inventor of the present invention has invented the present yogurt by utilizing the effect which is considered to be produced as a by-product from the environmental adaptability of microorganisms.

Specific examples of the genus Bifidus used in the present invention include B. bifidum,
B. longum, B. infantis, B. brev
e), A. dresses
is), catenulatum (B. catenulatu)
m), Pseudocatenulatum (B. pseudoc)
atenulatum, B. dent
ium), B. angulatu
m), B. gallicum, B. inopinatum, B. denticolens, B. pseudolongum, B. cuniculi, B. cholineum.
erinum), animalis (B. animali)
s), thermo film (B. thermophilu
m), Bohm, B.mag
unum), B. pullorum, B. suis, B. galli
narum), B. ruminan
Tium), Melissicum (B. merycicum),
Circulare (B. saeculare), minimum (B. minimum), subtil (B. subtil)
e), coryneform (B. coryneform)
e), Asteroides, B. indicum and the like. Of these, B. bifidum,
B. longum, B. infantis, B. brev
e), or B. animalis. One type of Bifidobacterium may be selected and used, or two or more types may be used in combination.

Hereinafter, for the sake of simplicity, the genus Bifidobacteria will be referred to as A
Lactococcus lactis subspecies lactis and Lactococcus lactis
Subspecies cremoris is called B group.

Bacteria A and B are added to the raw material which has been subjected to the steps of mixing, homogenizing, sterilizing and cooling as described above. The ratio of the A bacterium and B bacterium group is preferably about A bacterium: B bacterium group = 1: 1 to 1: 2. Specifically, for the mix
It is preferable to add about 1% to 2% of A bacterium and about 2% of B bacterium group. In addition, the total amount of the mixed bacteria of the A and B bacteria groups is preferably about 3% to 4% with respect to the total amount of the mix.

Fermentation temperature is 3 which is suitable for the growth of A bacterium.
0 to 40 ℃, moderate temperature of the mesophilic bacteria group B (20 ℃
-30 ° C). Particularly, the temperature is preferably 36 ° C to 38 ° C. Further, the pH of fermentation is 4.0 to 5.
The process ends when the number reaches 5, preferably 4.6 to 4.8. The fermentation time is preferably 5 hours to 10 hours, particularly preferably 7 hours to 9 hours.

FIG. 2 is a diagram showing an example of pH change with time of a mixture obtained by adding only A bacterium to milk and fermenting at 38 ° C. As shown in the figure, the pH gradually decreases with time. The pH was 6.20 after 11 hours had passed from the start of fermentation, and the mixture was in a state of being slightly coagulated.

On the other hand, FIG. 3 shows that only B bacteria group was added to milk.
It is a figure showing an example of pH change over time of a mixture fermented at 8 ° C. As shown in the figure, the pH changes rapidly with the passage of time. The pH after 5 hours from the start of fermentation is 5.14, and the mixture is in a solidified state. However, yogurt obtained only by using the B bacteria group is inferior in flavor and cannot obtain beneficial effects such as intestinal regulating action.

FIG. 4 is a diagram showing an example of pH change with time of a mixture obtained by adding bacteria A and B to milk and fermenting it at 38 ° C., which corresponds to yogurt according to the present invention. The pH change is almost the same as in FIG. 3, and after 6 hours from the start of fermentation, yogurt in a coagulated state is obtained at pH 5.25. Further, this yogurt has a beneficial effect represented by an intestinal regulating action together with an excellent flavor.

That is, according to the method of the present invention, the fermentation time can be shortened as compared with the case where only the bacterium A is added. Further, it is possible to obtain a flavor and an effect which cannot be obtained when only the B bacterium group is added. As described above, according to the method of the present invention, it is possible to produce yogurt having excellent characteristics even with a combination of lactic acid bacteria that has not been conventionally used because it has no symbiotic relationship.

After completion of fermentation, the curd is obtained by cooling to 5 ° C to 8 ° C. In the case of producing a stirred yogurt, the curd is further crushed and liquefied.

The method for producing yogurt according to the present invention can be applied to both static yogurt and stirred yogurt.

According to the present invention, Lactococcus lactis subspecies
It has been found that the combined use of Lactis, Lactococcus lactis subspecies cremoris and Bifidobacterium can produce yogurt even with a combination of lactic acid bacteria that are not symbiotic.

Further, according to the present invention, by using, as a lactic acid bacterium starter, a combination of Lactococcus lactis subspecies lactis, Lactococcus lactis subspecies cremoris and Bifidobacterium genus It was found that yogurt having a refreshing fragrance and flavor as well as an intestinal regulating action such as immediate improvement of constipation can be produced.

Example 1. The sterilization treatment was performed by heating and holding the raw milk at 85 ° C. for 15 minutes. Then, after cooling to 38 ° C., a mixture of Lactococcus lactis subspecies lactis and Lactococcus lactis subspecies cremoris (Rhodia, trade name EZAL M) was added to the raw milk.
R014) 2%, bifidobacteria (Rhody (Rho
dia), trade name EZALBL) was added at 1%.
Then, the mixture containing lactic acid bacteria was poured into the container, transferred to a fermenter, and fermented at a temperature of 38 ° C. for 8 hours. After fermentation,
A stationary yogurt was obtained by cooling to 5 ° C to 8 ° C.

Example 2. The sterilization treatment was performed by heating and holding the raw milk at 85 ° C. for 15 minutes. Then, after cooling to 38 ° C., a mixture of Lactococcus lactis subspecies lactis and Lactococcus lactis subspecies cremoris (Rhodia, trade name EZAL M) was added to the raw milk.
R014) 2%, bifidobacteria (Rhody (Rho
dia), trade name EZALBL) was added at 1%.
Then, the mixture containing lactic acid bacteria was poured into the container, transferred to a fermenter, and fermented at a temperature of 38 ° C. for 8 hours. After fermentation,
Cooled to 5-8 ° C. Subsequently, the pudding-shaped yogurt (curd) was crushed and then liquefied with a homogenizer to obtain a sugar-free drink yogurt as a stirring yogurt.

Example 3. 5% sugar was added to raw milk, mixed, and stirred to equalize. Then, the mixture was sterilized by heating and holding the mixture at 85 ° C. for 15 minutes. Then, after cooling the mixture to 38 ° C., to the raw milk,
A mixture of Lactococcus lactis subspecies lactis and Lactococcus lactis subspecies cremoris (Rhodia, trade name EZAL MR014), 2%, Bifidobacteria (Rhodia), product Name EZAL
BL) was added at 1%. Then, the mixture containing lactic acid bacteria was poured into the container, transferred to a fermenter, and fermented at a temperature of 38 ° C. for 8 hours. After the fermentation was completed, it was cooled to 5 ° C to 8 ° C. Subsequently, the pudding-shaped yogurt (curd) was crushed and then liquefied with a homogenizer to obtain a sugar-type drink yogurt as a stirring yogurt.

The types of bacteria contained in the yogurt obtained in Examples 2 and 3 and the viable cell count were analyzed.

Colonies appearing on a plate medium for measuring the number of viable cells were picked by type to obtain single colony isolation.
Ion) was repeated to produce a purified strain as BCGB.
DNA was extracted by the method. Using 1 μl of this as a template, primer A (5′-CCGCAGCCAA-
3 ') and primer B (5'-AACGCGCAA)
The electrophoretic patterns of the growth products were compared by the RAPD method using C-3 ') to narrow down the strains.

Also, from 1.0 ml of yogurt to BCGB
Using the DNA directly extracted by the method as a template, the bacterial species was identified by the PCR method using the bacterial species-specific primers. As a species-specific primer, animalis (B. animalis, BLAC), bifidum (B. bifidum, BiBIF), breve (B.
breve, BiBRE), longum (B. longgu
m, BiLON), Acidophilus (L. acidop
hilus, LbA), L. casei, Lb
C), L. delbrueckii, L
bD), gasserie (L. gasseri, LbG),
Helveticus (LbH),
Johnsonii (L. johnsonii, LbJ), plantarum (L. plantarum, LbP), rhamnose (L. rhamnosus, LbR), Lactococcus lactis subspecies cremoris (Lactococcus lactis subs).
p. cremoris, LcC), Lactococcus
Lactis Subspecies Lactis
ococcus lactis subsp. lac
tis, LcL) and Streptococcus thermop
hilus, ST) were used in total.

DNA analysis revealed that the yogurts obtained in Examples 2 and 3 were BLAC and LcC.
Bacterial species-specific amplification was observed with three types of primers, LcL and LcL, and the presence of three species of animalis, Lactococcus lactis subspecies cremoris and Lactococcus lactis subspecies lactis was confirmed.

By the culture method, four types of strains having different electrophoretic patterns were isolated from the yogurts obtained in Examples 2 and 3. FIG. 5 shows the result of strain identification by PARD of the isolates. In the figure, lane numbers are M for DNA size marker, 1 for Y50178 isolate, 2 for Y50179 isolate, 3 for Y50180 isolate, and 4 for Y501.
Representing 81 isolates.

FIG. 6 is a diagram showing the results of bacterial species identification by the PCR method. In the figure, Lactococcus lactis subspecies cremoris (Lactococc
uslactis subsp. cremoris,
LcC, Lactococcus lactis subspecies lactis (Lactococcus lact)
is subsp. Lactis, LcL) and animalis (B. animalis, BLAC) strain-specific bands were observed.

In FIG. 6, the lane numbers are M for DNA size marker and 5 for Lactococcus.
Ractis Subspecies Ractis, 6 for Y5
0178 isolates, 7 is Y50179 isolates, 8 is Y
50180 isolates, 9 Lactococcus lactis subspecies cremoris, 10 animalis,
And 11 represents the isolate of Y50181. From these results, isolate Y50178 was identified as Lactococcus lactis subspecies cremoris and Y501.
79 isolates and Y50180 with Lactococcus lactis subspecies lactis, Y501
81 were identified as animalis, respectively.

1 ml of yogurt was collected, and 0.1 ml of 10-fold serial dilution was added to TOS agar medium, Acidic.
-MRS agar (pH 5.4) and BCP-added plate count agar were smeared. The TOS agar medium and the Acidic-MRS agar medium (pH 5.4) were anaerobically cultured at 37 ° C for 3 days. Also, BCP-added plate count agar was aerobically aerated at 37 ° C for 3 days and 26 ° C.
Each was cultured for 7 days. The viable cell count per 1 ml of the sample after culturing was measured.

Table 1 shows the number of viable bacteria contained in 1 ml of the yogurt obtained in Examples 2 and 3. The viable cell counts shown in Table 1 are TOS agar medium for Animalis bacterium, Lactococcus lactis subspecies cremoris and Lactococcus lactis.
For Subspecies lactis, BCP-added plate count agar medium (cultured at 26 ° C.) was used for calculation.

[0053]

[Table 1]

As can be seen from Table 1, in the yogurt produced by the present invention, unlike conventional yogurt, a large amount of animalis, which belongs to the genus Bifidobacteria, was found to exist. It was also found that there are particularly many Lactococcus lactis subspecies lactis.

[0055]

According to the present invention, as a lactic acid bacterium starter, Lactococcus lactis subspecies
It has been found that the combined use of Lactis, Lactococcus lactis subspecies cremoris and Bifidobacterium can produce yogurt even with a combination of lactic acid bacteria that are not symbiotic.

Further, according to the present invention, by using a combination of Lactococcus lactis subspecies lactis, Lactococcus lactis subspecies cremoris and bifidobacteria as a lactic acid bacterium starter, conventional yogurt is used. It was found that yogurt having a refreshing fragrance and flavor as well as an intestinal regulating action such as immediate improvement of constipation can be produced.

[Brief description of drawings]

FIG. 1 is a diagram showing an example of a process for producing yogurt according to the present invention.

FIG. 2 is a diagram showing an example of a pH change with time of yogurt obtained by adding only bacteria A to milk.

FIG. 3 is a view showing an example of pH change with time of yogurt obtained by adding only B bacteria group to milk.

FIG. 4 is a graph of p of yogurt according to the invention with respect to time.
It is a figure showing an example of H change.

FIG. 5 is a diagram showing the results of strain identification by PARD of isolated strains.

FIG. 6 is a diagram showing the results of bacterial species identification by the PCR method.

[Explanation of symbols]

1,6 Y50178 isolate, 2,7 Y5017
9 isolates, 3,8Y50180 isolates, 4,1
1 Y50181 isolates, 5 Lactococcus lactis subspecies lactis, 9 Lactococcus lactis subspecies cremoris, 10 animalis.

─────────────────────────────────────────────────── ─── Continuation of the front page (58) Fields surveyed (Int.Cl. ⁷ , DB name) A23C 9/123-9/13 BIOSIS (DIALOG) JISST file (JOIS)

Claims (4)

(57) [Claims] 1. Lactococcus lactis (Lactococcus la)
ctis subsp. lactis), Lactococcus lactis subspecies cremoris (L
actococcus lactis subsp.
cremoris) and bifidobacteria (bifid)
and yogurt characterized by containing an bacterium. 2. The animal species (B.
animalis) yogurt according to claim 1. 3. A method for producing yogurt using lactic acid bacteria, wherein said lactic acid bacteria are Lactococcus lactis subspecies lactis (Lactococcus la).
ctis subsp. lactis), Lactococcus lactis subspecies cremoris (L
actococcus lactis subsp.
cremoris) and bifidobacteria (bifid)
and a method for producing yogurt, which is a mixed bacterium comprising bacteria. 4. The Bifidobacterium genus is B. bifidum or longum (B. longgu).
m), B. infantis, B. breve and animalis (B. an).
The method for producing yogurt according to claim 3, wherein the yogurt is at least one selected from the group consisting of imalis).