KR20220056047A - optimized medium for culturing Lactobacillus plantarum strain and method for culturing Lactobacillus plantarum strain using the same - Google Patents

Abstract

The present invention relates to a medium for culturing a Lactobacillus plantarum strain containing maltose as a carbon source, and a yeast extract and soytone as a nitrogen source, and a method for culturing a Lactobacillus plantarum strain using the same. According to the present invention, culturing efficiency can be increased.

Description

Optimized medium for culturing Lactobacillus plantarum strain and culturing method of Lactobacillus plantarum strain using same

The present invention relates to an optimized medium for culturing a Lactobacillus plantarum strain and a method for culturing a Lactobacillus plantarum strain using the same.

Probiotics refer to living microorganisms that, when consumed in appropriate amounts, improve the intestinal microflora and have a beneficial effect on the host. As the consumption of probiotic products increases, technology to apply lactic acid bacteria to food, mass production of lactic acid bacteria, and production optimization research are also being actively conducted. The genus Lactobacillus , along with the genus Bifidobacterium , is considered a probiotic and is one of the microorganisms with difficult growth conditions. For high growth of Lactobacillus, a medium containing nutrients such as amino acids, peptides, vitamins, and nucleic acids is required. In addition to medium components, culture conditions such as pH and temperature also significantly affect the growth of Lactobacillus. Therefore, it is industrially important to find the optimal culture conditions according to the strain.

One-factor-at-a-time (OFAT) experiment is a simple experimental design method in which only one factor is changed while other factors are fixed at a constant level, so it has been widely used in various optimization experiments. However, OFAT requires a considerable amount of time and labor, and has a disadvantage in that it is difficult to determine the effect of factors. On the other hand, the response surface methodology (RSM) is a statistical analysis method that includes a full factor batch method and regression analysis. Since the surface formed by the response variable is statistically analyzed, the amount of response according to the change of the regression variable can be predicted. In addition, it is possible to estimate the values of the regression variables that are the minimum, maximum, and optimum of the response amount. Among several experimental design methods, central composite design (CCD) is being used as an effective method when establishing optimal conditions through RSM.

The present invention provides a medium for optimally culturing a Lactobacillus plantarum strain, as a carbon source, maltose; And as a nitrogen source, including yeast extract (yeast extract) and soytone (soytone), Lactobacillus plantarum ( Lactobacillus plantarum ) To provide a culture medium and the like of the strain.

However, the technical problem to be achieved by the present invention is not limited to the above-mentioned problems, and other problems not mentioned will be clearly understood by those skilled in the art from the following description.

The present invention as a carbon source, maltose (maltose); And as a nitrogen source, including yeast extract (yeast extract) and soytone (soytone), Lactobacillus plantarum ( Lactobacillus plantarum ) Provides a culture medium for the strain.

The concentration of the maltose is 3.0 to 3.5% (w/v), and the concentrations of the yeast extract and soytone are 2.7 to 3.2% (w/v) and 3.7 to 4.2%, respectively. (w/v).

The medium may further include a trace element selected from the group consisting of MnSO ₄ , MgSO ₄ , K ₂ HPO ₄ , KH ₂ PO ₄ and NaCl at a concentration of 0.001 to 1.0% (w/v).

The Lactobacillus plantarum ( Lactobacillus plantarum ) strain may be Lactobacillus plantarum ( Lactobacillus plantarum ) 200655 strain (KCCM 12204P).

In one embodiment of the present invention, the medium, Lactobacillus plantarum ( Lactobacillus plantarum ) comprising the step of inoculating and culturing the strain, Lactobacillus plantarum ( Lactobacillus plantarum ) Provides a method of culturing the strain.

The culture may be performed at an initial pH of 4.5 to 9.0 and a temperature of 25 to 35°C.

Lactobacillus plantarum according to the present invention The culture medium of the strain is a carbon source, maltose (maltose); And as a nitrogen source, the bar characterized in that it comprises a yeast extract and soytone (soytone), Lactobacillus plantarum ( Lactobacillus plantarum ) strain (in particular, Lactobacillus plantarum ) 200655 strain (KCCM 12204P)) when inoculated and cultured, since it can represent an increased culture efficiency than that in a commercial medium such as MRS medium used in the past, industrial Lactobacillus plantarum ( Lactobacillus plantarum ) strain It can be effectively used in culture.

1 is a graph comparing the difference in dry cell mass of Lactobacillus plantarum 200655 strain according to various carbon sources in the medium.
2 is a graph comparing the difference in dry cell mass of Lactobacillus plantarum 200655 strain according to various nitrogen sources in the medium.
3 is a graph comparing the difference in dry cell mass of Lactobacillus plantarum 200655 strain according to various trace elements of the medium.
4 is a contour diagram showing the interaction of maltose, yeast extract, and soytone with respect to the dry cell mass of the Lactobacillus plantarum 200655 strain [(A) maltose and soytone ( interaction of soytone, (B) interaction of maltose and yeast extract, (C) interaction of yeast extract and soytone].
5 is a three-dimensional response surface diagram showing the interaction of maltose, yeast extract, and soytone with respect to the dry cell mass of Lactobacillus plantarum 200655 strain [(A) maltose and interaction of yeast extract, (B) interaction of maltose and soytone, (C) interaction of yeast extract and soytone].
6 is a graph comparing the dry cell mass of Lactobacillus plantarum 200655 strain in RSM optimized medium and MRS medium.
7 is a graph comparing the difference in dry cell mass of Lactobacillus plantarum 200655 strain according to the initial pH during culture.
8 is a graph comparing the difference in dry cell mass of Lactobacillus plantarum 200655 strain according to temperature during culture.

The present inventors predicted the optimal concentration of factors affecting the increase in dry cell mass by using the response surface analysis method while studying the medium for optimally culturing the Lactobacillus plantarum strain. In addition, the suitability of the experimental model was confirmed through statistical analysis, and the value predicted by the experimental model was verified by experiment, confirming that the reliability of the experimental model was high. After optimizing the medium, by optimizing the culture conditions of the Lactobacillus plantarum strain by checking the culture conditions according to the initial pH and temperature, the present invention was completed.

First, the terminology of the present invention will be described.

The term "dry cell weight" used in the present invention may be used in the same meaning as the dry cell weight of the microbial strain cultured in the medium. Even if the number of viable microorganisms cultured in the medium is similar, if there is a difference in "dry cell mass", it can be understood that the size grown per single microorganism is different, which may indicate a difference in dry weight. That is, even if the number of viable cells is shown at a similar or the same level in the culture results of microbial strains cultured in different media, if there is a difference in the "dry cell mass", the culture efficiency is better than under the culture condition in which the cell mass appears at a high level. can be understood

In addition, the term "culture efficiency" used in the present invention means the maximum cell yield compared to the culture time. The time that can represent the maximum amount of dry cells during the culture time is set as the optimal culture time, and the maximum cell yield compared to the culture time is obtained by calculating the dry cell mass according to the optimal culture time as a ratio. This can be expressed as culture efficiency.

Hereinafter, the present invention will be described in detail.

The present invention as a carbon source, maltose (maltose); And as a nitrogen source, including yeast extract (yeast extract) and soytone (soytone), Lactobacillus plantarum ( Lactobacillus plantarum ) Provides a culture medium for the strain.

The medium according to the present invention is for optimally culturing the Lactobacillus plantarum strain, and as a carbon source, maltose; And as a nitrogen source, it includes yeast extract and soytone, and it is characterized in that the optimal concentration is derived based on the result of the reaction surface analysis method.

First, the medium contains maltose as a carbon source, and the concentration of maltose is preferably 3.0 to 3.5% (w/v), and the concentration of maltose is 3.0 to 3.3%. (w/v) is more preferable, but is not limited thereto.

In addition, the medium includes a yeast extract and soytone as a nitrogen source, and the concentrations of the yeast extract and soytone are 2.7 to 3.2% (w/v), respectively. and 3.7 to 4.2% (w/v), and the concentrations of the yeast extract and soytone are 2.9 to 3.1% (w/v) and 3.85 to 4.05% (w/v), respectively. ) is more preferable, but is not limited thereto.

In addition, the medium may further include a trace element selected from the group consisting of MnSO ₄ , MgSO ₄ , K ₂ HPO ₄ , KH ₂ PO ₄ and NaCl at a concentration of 0.001 to 1.0% (w/v), MnSO ₄ Or MgSO ₄ It is preferred to further include 0.05 to 0.5% (w / v) concentration, but is not limited thereto.

In addition, the medium further contains sodium acetate 0.3 to 0.8% (w/v), K ₂ HPO ₄ 0.1 to 0.5% (w/v) and tween 80 0.05 to 0.2% (w/v) can do.

On the other hand, the Lactobacillus plantarum ( Lactobacillus plantarum ) strain may be Lactobacillus plantarum ( Lactobacillus plantarum ) 200655 strain (KCCM 12204P).

Specifically, the strain (KCCM 12204P) was deposited with the Korea Microorganism Conservation Center on January 17, 2018 and was given an accession number KCCM 12204P. The strain (KCCM 12204P) is a subspecies of Lactobacillus plantarum, which is a Gram-positive bacillus and a slightly aerobic strain, and is one of the most widely distributed lactic acid bacteria in nature. Lactobacillus plantarum is a typical kimchi lactic acid bacterium that undergoes normal lactic acid fermentation and produces lactic acid by fermenting arabinose, glucose, fructose, galactose, maltose, sucrose, and dextran. It is mainly isolated from dairy products, pickles, pickled vegetables such as kimchi, tomatoes, etc., especially in the late stage of kimchi fermentation, when kimchi is fermented and sour, it mainly grows and dominates, inhibiting the growth of other species. It is known to have excellent bile resistance. In particular, the strain (KCCM 12204P) has an immune enhancing effect, degenerative neurological disease prevention or treatment efficacy, and by optimally culturing the strain (KCCM 12204P) using the medium according to the present invention, the efficacy can be maximized. there is.

In addition, the present invention, Lactobacillus plantarum ( Lactobacillus plantarum ) in the medium, comprising the step of inoculating and culturing the strain, Lactobacillus plantarum ( Lactobacillus plantarum ) Provides a method of culturing the strain.

The culture may be performed by any one of a culture method of stationary culture, fed-batch culture, and batch culture, and specifically, the culture is Lactobacillus plantarum It is carried out at a temperature suitable for growth. Preferably, the initial pH of 4.5 to 9.0 (preferably 6.5 to 7.5) and 25 to 35 ℃ (preferably, it can be carried out at a temperature of 27 to 32 ℃. On the other hand, outside the initial pH range or temperature range In this case, there is a problem in that the culture efficiency is lowered.

As discussed above, the culture medium of the Lactobacillus plantarum strain according to the present invention is a carbon source, including maltose; And as a nitrogen source, the bar characterized in that it comprises a yeast extract and soytone (soytone), Lactobacillus plantarum ( Lactobacillus plantarum ) strain (in particular, Lactobacillus plantarum ) 200655 strain (KCCM 12204P)) when inoculated and cultured, since it can represent an increased culture efficiency than that in a commercial medium such as MRS medium used in the past, industrial Lactobacillus plantarum ( Lactobacillus plantarum ) strain It can be effectively used in culture.

Hereinafter, preferred examples are presented to help the understanding of the present invention. However, the following examples are only provided for easier understanding of the present invention, and the contents of the present invention are not limited by the following examples.

<Example>

Example 1: Lactobacillus plantarum 200655 Preparation of strain and measurement of dry cell mass

As the Lactobacillus plantarum strain in the present invention, Lactobacillus plantarum 200655 strain (KCCM 12204P) was used. Lactobacillus plantarum 200655 strain is known as a lactic acid bacterium derived from kimchi, and was provided and used by the Korea Microbial Conservation Center. Lactobacillus plantarum 200655 strain was inoculated in MRS medium, and subcultured a total of 3 times at intervals of 12 hours while stationary culture at 37 ℃. After adjusting the concentration of the strain to be 0.5±0.05 at absorbance at 600 nm, it was transferred to the main culture and 1% (w/v) of Lactobacillus plantarum 200655 strain was inoculated and cultured at 32° C. for 24 hours.

Then, 50 mL of the culture medium was collected, centrifuged at 4,000Х g, 4° C. for 15 minutes, and then washed twice with sterile distilled water. After drying until a constant weight was reached in a dryer maintained at 80°C, the mass was measured.

Example 2: Selection of carbon source, nitrogen source and trace element for cell growth

In order to optimize the culture medium for the Lactobacillus plantarum 200655 strain, the carbon source, nitrogen source, and trace element showing the highest influence on cell growth were selected. The basal medium consists of: 5 g/L sodium acetate (Sigma, USA), 2 g/LK ₂ HPO ₄ (Samchun, Korea) 0.1 g/L MgSO ₄ 7 H ₂ O (Shinyo pure chemicals Co, Japan) ), 0.05 g/L MnSO ₄ .H ₂ O (Samchun, Korea) and 1 g/L tween 80 (Yakuri pure chemicals Co. Ltd, Japan). 100 mL of the medium was prepared and used for the experiment. After preparing the medium, the initial pH was adjusted to 6.5 using 1 M HCl and 1 M NaCl, and then sterilized at 121° C. for 15 minutes by autoclave.

The type of carbon source was selected after examining the carbon source availability of the strain using the API 50 CHL medium kit. Carbon sources include glucose (glucose, Sigma, USA), sucrose (sucrose, Samchun, Korea), maltose (maltose, Junsei, Japan), fructose (fructose, Merck, German), lactose (lactose, Difco, USA), Galactose (galactose, Difco, USA) was used. A medium was prepared by adding 2% (w/v) of a carbon source to a basal medium to which 0.5% (w/v) of yeast extract (Difco, USA) was added. 1 is shown.

As shown in FIG. 1 , the dry cell mass in the medium using maltose as a carbon source gave the highest value at 1.394±0.067 g/L, followed by sucrose, galactose, and lactose. (lactose), glucose (glucose), fructose (fructose) 1.349±0.062 g/L and 0.731±0.057 g/L, respectively. Dry cell masses of 0.501±0.086 g/L, 0.291±0.172 g/L, and 0.084±0.043 g/L were measured. Therefore, it was determined that the carbon source in the optimized medium for culture of the Lactobacillus plantarum 200655 strain was maltose.

As nitrogen sources, tryptone (tryptone, Difco, USA), soytone (soytone, Difco, USA), peptone (peptone, Difco, USA), yeast extract (Difco, USA), beef extract (beef extract, Difco) , USA) and malt extract (Difco, USA) were used. A medium was prepared by adding 1% (w/v) of a nitrogen source to the basal medium to which 2% (w/v) of maltose was added, and the results of measuring the dry cell mass after stationary culture are shown in FIG. 2 .

As shown in Figure 2, the dry cell mass in the medium using yeast extract and soytone as a nitrogen source was 1.551 ± 0.106 g / L and 1.069 ± 0.022 g / L, respectively. Then, tryptone, beef extract, peptone, and malt extract were followed in the order of 0.751±0.063 g/L, 0.682±0.064 g/L, and 0.577±0.060 g/L, respectively. L, the dry cell mass of 0.181±0.075 g/L was measured. Therefore, it was determined that the nitrogen source in the optimized medium for culture of Lactobacillus plantarum 200655 strain was yeast extract or soytone.

As trace elements, MgSO ₄ , MnSO ₄ , K ₂ HPO ₄ , KH ₂ PO ₄ , and NaCl were used. A medium was prepared by adding 0.1% (w/v) of trace elements to a basal medium to which 2% (w/v) of maltose and 1% (w/v) of yeast extract were added, and cultured stationary. The results of measuring the dry cell mass afterward are shown in FIG. 3 .

As shown in FIG. 3 , the dry cell mass in the medium using MnSO ₄ as a trace element gave a high value of 0.807±0.032 g/L, and other trace elements MgSO ₄ , K ₂ HPO ₄ , KH ₂ PO ₄ , The values were 1.77 times, 1.69 times, 1.68 times, and 1.74 times higher than those of NaCl. Therefore, it was determined that the trace element in the culture optimized medium for the Lactobacillus plantarum 200655 strain is MnSO ₄ .

Example 3: Using Response Surface Analysis Lactobacillus plantarum Optimization of the culture medium for the 200655 strain

In order to determine the optimal concentration of the carbon source, nitrogen source and trace element selected in Example 2, a response surface methodology (RSM) was performed using a central composite design (CCD). In Example 2, maltose, yeast extract, and MnSO ₄ were selected as factors with high influence, but MnSO ₄ had relatively low influence compared to other factors, and yeast extract required by the strain (yeast). Since the amount of extract) was significant, the experiment was conducted by changing MnSO ₄ to soytone. As independent variables, maltose (g/L, X ₁ ), yeast extract (g/L, X ₂ ), and soytone (g/L, X ₃ ) were three factors - It was coded in five levels as α, -1, 0, 1, α. The experimental design was set to 20 experimental spheres including 8 vertices, 6 axial points, and 6 central points according to the central synthesis programming method. The dependent variable (Y) according to the independent variable was the dry cell mass, and the medium was prepared and tested according to the set conditions. The regression analysis model equation is as follows.

와

는 독립변수,

은 절편,

는 회계계수이다. Minitab 18 program을 이용하여 실험 계획 수립과 통계 분석하였고 회귀방정식 모델을 도출하였다. 이 방정식을 바탕으로 최대 균체량 생산이 가능한 각 요인들의 최적 농도를 결정하였으며, 실험 모델의 유의성 또한 확인하였다. Y is the dependent variable,

Wow

is the independent variable,

silver intercept,

is an accounting coefficient. Experimental planning and statistical analysis were performed using Minitab 18 program, and a regression equation model was derived. Based on this equation, the optimal concentration of each factor capable of producing the maximum cell mass was determined, and the significance of the experimental model was also confirmed.

Response surface analysis was performed on the previously selected maltose, yeast extract, and MnSO ₄ three factors. At this time, the central point of each factor was set to 25 g/L, 55 g/L, and 0.15 g/L, and the experiment was conducted using the central synthesis programming method of 20 experimental groups. As a result of the experiment, it was confirmed that each factor had a positive effect on the dry cell mass. However, the p-value of MnSO ₄ was not significant with a p- value of 0.05 or more, and the coefficient of influence was also measured to be very low as 0.0083, so it was excluded. In addition, in the case of yeast extract, since a high concentration of 71.33 g/L or more was required, the experiment was modified and conducted because it did not meet the industrially easy optimal medium production goal. Therefore, MnSO ₄ was added as a basic medium component at a concentration of 0.05 g/L, and soytone was added to supplement the nitrogen source.

Maltose (X ₁ ), yeast extract (X ₂ ), soytone (X ₃ ) three factors were used as independent variables to establish an experimental plan using the central synthetic programming method. For each independent variable, a preliminary experiment was conducted to set the central point at the concentrations of 25 g/L, 25 g/L, and 30 g/L, respectively. Based on this, five levels were coded: -1.68, -1, 0, 1, and 1.68. The reaction value (Y, dry cell mass) was measured by executing 20 experimental groups including 6 center points (Table 1).

independent variable sign
(unit) Encrypted Variable Level - α One 0 One α Maltose X ₁ (g/L) 8.18 15 25 35 41.82 yeast extract X ₂ (g/L) 8.18 15 25 35 41.82 Soytone X ₃ (g/L) 4.77 15 30 45 55.23 number variable Y (g/L) X ₁ x ₂ X ₃ One -One -One -One 2.484 2 One -One -One 3.216 3 -One One -One 2.620 4 One One -One 3.486 5 -One -One One 2.896 6 One -One One 3.714 7 -One One One 3.196 8 One One One 3.892 9 -1.682 0 0 2.348 10 1.682 0 0 3.570 11 0 -1.682 0 3.354 12 0 1.682 0 3.650 13 0 0 -1.682 3.232 14 0 0 1.682 3.546 15 0 0 0 3.846 16 0 0 0 3.684 17 0 0 0 3.610 18 0 0 0 3.794 19 0 0 0 3.810 20 0 0 0 3.728

The data were analyzed using the Minitab 18 program, and the regression equation for the dry cell mass was as follows:

의 값 또한 0.9676으로 1에 가까워 모형의 적합함을 확인하였다. 회귀방정식의 독립변수와 종속변수의 관계의 적합성을 확인하는 적합성 결여의 p-value는 0.1774로 나타났다. 이는 유의성 판단 기준인 0.05보다 높은 값으로 적합성 결여가 유의하지 않음을 의미하며 이는 회귀모델을 설명하는데 적합함을 의미한다(표 2).As a result of the analysis of variance, the p -value of the model showed a high significance of 0.0001 or less, and the coefficient of determination

The value of was also 0.9676, close to 1, confirming the fit of the model. The p -value of the lack of fit, which confirms the fit of the relationship between the independent and dependent variables of the regression equation, was 0.1774. This is a value higher than 0.05, the criterion for determining significance, meaning that the lack of fit is not significant, which means that it is suitable for explaining the regression model (Table 2).

sauce coefficient prediction DF
(Degree of freedom) Adj SS
(Adjusted sum of square) Adj MS
(Adjusted mean square) F-value p-value Model 3.7475 9 3.9906 0.4434 33.22 <0.0001 ^*** X ₁ 0.3784 One 1.9550 1.9550 146.46 <0.0001 ^*** x ₂ 0.1012 One 0.1398 0.1398 10.47 0.0089 ^*** X ₃ 0.1772 One 0.4289 0.4289 32.13 0.0002 ^*** X ₁ X ₂ 0.0015 One 0.0000 0.0000 0.00 0.9714 X ₁ X ₃ -0.0105 One 0.0009 0.0009 0.07 0.8024 X ₂ X ₃ 0.0090 One 0.0006 0.0006 0.05 0.8301 X ₁ ² -0.2922 One 1.2306 1.2306 92.18 <0.0001 ^*** X ₂ ² -0.1002 One 0.1448 0.1448 10.85 0.0081 ^*** X ₃ ² -0.1402 One 0.2832 0.2832 21.22 0.0010 ^*** residue 10 0.1335 lack of fit 5 0.0944 2.42 0.1774 Pure error 5 0.0391 total 19 4.1241

In order to examine the interaction according to each independent variable with respect to the dry cell mass, one independent variable was fixed at the optimal point, and the contour and 3D response surface diagrams were examined for the remaining two independent variables, and the results are shown in FIG. 4 and 5, respectively.

Through response surface analysis, the maximum dry cell mass of Lactobacillus plantarum 200655 strain was predicted to be 3.9514 g/L, and the optimal concentrations of each variable were 31.6252 g/L of maltose, 29.9265 g/L of yeast extract, and 29.9265 g/L of soy. 39.4284 g/L of soytone.

In order to verify the dry cell mass in the RSM-optimized medium, the medium was prepared according to the optimum concentration of each medium component. Lactobacillus plantarum 200655 was inoculated with 1% (w/v) and cultured at 32° C. for 24 hours, and then the dry cell mass was measured. As a control for comparison with the optimized medium, MRS medium commonly used for culturing lactic acid bacteria was used. In addition to the medium components, the experiment was conducted with the same temperature, incubation time, and inoculation ratio, and the results are shown in FIG. 6 .

As shown in FIG. 6 , as a result of culturing in RSM-optimized medium, the dry cell mass was 3.8453±0.0595 g/L. This is a value similar to 3.9514 g/L predicted through the experimental model, and it is confirmed that the predicted value of the experimental model is valid and reliable. In addition, compared to the dry cell mass of 2.4287±0.0420 g/L in the control MRS medium, it is confirmed that the cell production increased by 58.33% (w/v).

Example 3: Lactobacillus plantarum Optimization of culture conditions of the 200655 strain

In the culture of the Lactobacillus plantarum 200655 strain, an RSM-optimized medium was prepared to establish optimal production conditions according to the initial pH and temperature, and the experiment was conducted.

After setting the initial pH of the RSM-optimized medium to pH 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 1% (w/v) of Lactobacillus plantarum 200655 strain was inoculated at 32 ° C. After stationary culture for 24 hours, the dry cell mass was measured, and the results are shown in FIG. 7 . 1M HCl and 1M NaCl were used for pH adjustment.

As shown in FIG. 7 , at the initial pH of 4.0, the dry cell mass was 1.124±0.119 g/L, showing the lowest value. From the initial pH of 4.5 to the initial pH of 7.0, the dry cell mass gradually increased, showing the highest value at pH 7.0 at 4.304±0.118 g/L. Thereafter, from the initial pH of 7.5 to the initial pH of 9.0, there was a slight decrease. Therefore, it is confirmed that the Lactobacillus plantarum 200655 strain can be optimally cultured at an initial pH of 4.5 to 9.0 (preferably, 6.5 to 7.5).

On the other hand, after adjusting the pH of the RSM-optimized medium to 7.0, the culture temperature was set to 22 ℃, 25 ℃, 27 ℃, 30 ℃, 32 ℃, 35 ℃, Lactobacillus plantarum 200655 strain 1% (w / v) inoculation After stationary culture at 32° C. for 24 hours, the dry cell mass was measured, and the results are shown in FIG. 8 .

As shown in FIG. 8 , the dry cell mass at 32° C., which is the existing temperature condition, showed 4.373±0.080 g/L, while the dry cell mass at 22° C. showed the lowest value at 3.156±0.048 g/L. From 22°C to 30°C, the dry cell mass gradually increased, showing the highest value at 30°C of 4.544±0.083 g/L. After that, it showed a slightly decreasing pattern from 30°C to 35°C. Therefore, it is confirmed that the Lactobacillus plantarum 200655 strain can be optimally cultured at 25°C to 35°C (preferably, 27°C to 32°C).

The description of the present invention described above is for illustration, and those of ordinary skill in the art to which the present invention pertains can understand that it can be easily modified into other specific forms without changing the technical spirit or essential features of the present invention. will be. Therefore, it should be understood that the embodiments described above are illustrative in all respects and not restrictive.

Korea Microorganism Conservation Center (Overseas) KCCM12204P 20180117

Claims (6)

As the carbon source, maltose; and
As a nitrogen source, including yeast extract (yeast extract) and soytone (soytone), Lactobacillus plantarum ( Lactobacillus plantarum ) Medium for culture of the strain.
The method of claim 1,
The concentration of the maltose is 3.0 to 3.5% (w/v), and the concentrations of the yeast extract and soytone are 2.7 to 3.2% (w/v) and 3.7 to 4.2%, respectively. (w / v), characterized in that, Lactobacillus plantarum ( Lactobacillus plantarum ) Medium for culture of the strain.
The method of claim 1,
The medium is MnSO ₄ , MgSO ₄ , K ₂ HPO ₄ , KH ₂ PO ₄ And a trace element selected from the group consisting of NaCl at a concentration of 0.001 to 1.0% (w / v) Lactobacillus, characterized in that it further comprises Plantarum ( Lactobacillus plantarum ) Medium for culture of the strain.
The method of claim 1,
The Lactobacillus plantarum ( Lactobacillus plantarum ) strain is Lactobacillus plantarum ( Lactobacillus plantarum ) 200655 strain (KCCM 12204P), characterized in that, Lactobacillus plantarum ( Lactobacillus plantarum ) Culture medium for the strain.
Claims 1 to 4 in the medium according to any one of claims 1 to 4, Lactobacillus plantarum ( Lactobacillus plantarum ) Inoculating and culturing the strain, Lactobacillus plantarum ( Lactobacillus plantarum ) A method of culturing a strain.
6. The method of claim 5,
The culture is characterized in that the initial pH of 4.5 to 9.0 and carried out at a temperature of 25 to 35 ℃, Lactobacillus plantarum ( Lactobacillus plantarum ) Culture method of the strain.

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