JPH08116872A - Culture base for lactobacillus starter and production of cheese using the base - Google Patents

Abstract

PURPOSE: To provide the subject new culture base containing milk as a nitrogen source and a carbon source, calcium carbonate as an agent for suppressing the depression of pH and a soluble Mg salt as an agent for promoting the proliferation of lactobacillus and capable of giving a lactobacillus starter having high and stabilized activity at a low cost. CONSTITUTION: This culture base for the preparation of a lactobacillus starter contains a milk (preferably skim milk or whey powder) as a nitrogen source and a carbon source. The culture base is incorporated with calcium carbonate or calcium phosphate in an amount of preferably >=1.5% for suppressing the lowering of pH caused by the lactic acid fermentation and with >=150mM of a soluble magnesium salt (preferably magnesium sulfate or magnesium chloride) to promote the proliferation of lactobacillus. The amount of the skim milk or the whey powder is preferably 3.5-5%.

Description

Detailed Description of the Invention

[0001]

FIELD OF THE INVENTION The present invention relates to a novel culture medium used for preparing a lactic acid bacterium starter for producing dairy products. More specifically, it relates to the composition of a culture medium for preparing a lactic acid bacteria starter used when producing a fermented milk product such as cheese, fermented milk, a lactic acid bacterium beverage, and a fermenting butter, and further to a method for producing cheese using this culture medium.

[0002]

Lactic acid fermentation is widely used in the production of food products. Especially in the dairy industry, there are many fermented milk products produced by lactic acid fermentation such as cheese, fermented milk, lactic acid bacterium drink, and fermented butter. In order to produce these fermented dairy products, usually, the lactic acid bacterium according to the use is inoculated into a culture medium, and the lactic acid bacterium culture obtained by culturing in advance is used as a lactic acid bacterium starter (hereinafter referred to as starter). The method of adding to the main raw material of and carrying out the main fermentation is generally adopted. As this culture medium, skim milk or reduced skim milk having a solid content of about 10% is usually used, and a component containing a growth factor such as yeast extract has been blended as necessary. In addition to such natural culture media, synthetic culture media are also used for research on lactic acid bacteria. For example, GYP medium (Uchimura et al., Lactic Acid Bacteria Experiment Manual, 50 pages, published by Asakura Shoten, 1992), LMB medium (Agricultural Chemistry Department, Faculty of Agriculture, University of Tokyo, Experimental Agricultural Chemistry Vol. 182, Asakura Shoten,
1981), M-17 medium (Appl. Microbiol., Vol. 2
9,807,1973), MRS medium (J. Appl. Bacteriol., Vol.
23,130-135,1960), Ericher medium (J. Dairy Sci., Vol.3
There are synthetic culture media such as 9,1611,1956). However, these synthetic culture media have not been used in actual production because their composition components are expensive, leading to an increase in production cost, and because a starter excellent in lactic acid fermentation ability cannot always be prepared.

Recently, a culture medium has been proposed for the purpose of imparting phage resistance and improving storage stability by adding specific components to skim milk for starters. For example, as the former example, JP-A-60-105489 discloses a medium in which orthophosphate is added to skim milk and an organic acid is added after boiling. In addition, JP-A-60-7523
Japanese Patent Publication No. 3 discloses a technique of adding an ester of oleic acid to skim milk and culturing. However, such additives also do not function to increase the vitality of the starter as disclosed in the present invention. Since skim milk or skim milk powder is a natural product, it is known that the quality is not constant and varies depending on the season. Therefore, due to the influence, the fermentation ability of the lactic acid bacterium starter prepared using them as a culture medium varies, which has been a problem in process control. Therefore, a culture medium capable of preparing a lactic acid bacterium starter having a higher and stable lactic acid fermentation ability has been desired.

[0004]

SUMMARY OF THE INVENTION There is a need to provide a culture medium suitable for producing a starter for use in producing fermented milk products such as cheese, fermented milk, lactic acid bacteria beverages and fermented butter. In the conventional culture medium, the lactic acid fermentation ability of the prepared starter was not stable, and a starter having a low lactic acid fermentation ability was often prepared. Such a starter is expressed as having low vitality, which means that the starter produces a small amount of lactic acid within a predetermined culture time. It has been pointed out that the use of a low-activity starter extends the production time of cheese and fermented milk, often causing product defects. An object of the present invention is to provide a composition of a culture medium for preparing a highly active and stable starter. It is also an object of the present invention to provide a cheaper culture medium and reduce the production cost of the starter. Furthermore, it is also an object of the present invention to shorten the time required for cheese production by using the starter provided by the present invention.

[0005]

The starter culture medium of the present invention uses milk, particularly skim milk powder or whey (whey) powder, as a nitrogen source and a carbon source. Although skim milk powder or skim milk has been conventionally used as a component of a culture medium for a starter, as described above, the quality of skim milk or skim milk powder varies depending on the season, and thus a starter containing these as a culture medium is used. The vitality of is also prone to instability. Therefore, in the present invention, skim milk powder or whey powder is used as an inexpensive nitrogen source and carbon source, and further, skim milk powder or whey powder may be 3% or more as the nitrogen source and carbon source of lactic acid bacteria. confirmed. Particularly preferably, it should be 3.5% or more, and coagulation occurs when the solid content concentration is 5% or more. In the following description of the present invention, skim milk powder or whey powder was used, but of course skim milk or whey (whey) can also be used,
When using such a liquid raw material, it is advisable to measure the milk solid content and dilute it so as to be in the range of 3.5 to 5%.

The skim milk powder or whey powder, which is a nitrogen source and a carbon source, can be used by simply dissolving it in water to a predetermined concentration. The product thus dissolved is referred to as reduced skim milk or reduced whey. Conventionally, the reduced skim milk or reduced whey was sterilized by heating, cooled, inoculated with a target lactic acid bacterium, and generally cultured overnight at 22 ° C. to prepare a starter. At this time, it is preferable to add about 0.1 to 1.0% of a component such as yeast extract, if necessary, for the growth of the lactic acid bacterium, but it is not necessary to add it. It has been known that an inorganic salt having a buffering capacity such as sodium may be added in an amount of about 0.5 to 2%.

[0007] However, one of the problems with the conventional culture medium is that the lactic acid fermentation ability of the lactic acid bacterium decreases as soon as the starter culture is completed, resulting in a less active starter. The cause of this is that the pH of the culture is lowered by the lactic acid produced by lactic acid bacteria. In order to solve such a problem, the addition of a salt having a pH buffering ability as described above has been performed, but it was not always effective in high-density culture of lactic acid bacteria such as a starter. Therefore, it has been empirically carried out to add a neutralizing agent such as calcium carbonate to the culture medium in advance. What is the optimum salt for maintaining the pH of the culture at an optimum value, or There have been no specific studies on how appropriate the added amount is. In the present invention, as shown in the following test examples, as a result of examining the optimum amount and the salt to be used, it was found that calcium carbonate or calcium phosphate maximizes the vitality of the starter. In the composition of the culture medium for starter of the present invention, this calcium carbonate or calcium phosphate is
1% or more, particularly preferably 1. 5% or more is contained. Calcium carbonate or calcium phosphate are insoluble fine particles and become calcium lactate along with the production of lactic acid.
The decrease of H is suppressed. If the culture medium of the present invention contains about 1% or more of calcium carbonate or calcium phosphate, its pH during fermentation is the optimum pH of lactic acid bacteria.
It can be maintained at ~ 6.5, and since it does not affect the production of lactic acid, the vitality of the starter is kept high.

Further, in order to enhance the vitality of the starter, the utilization of a lactic acid production promoting factor can be mentioned. Up to now, the utilization of the production promoting factor has been mainly to utilize natural ingredients such as yeast extract and meat extract which are the production promoting factors. However, the studies conducted by the present inventors have revealed that the presence of magnesium is particularly important in the case of a starter culture medium using milk protein. In the conventional culture of lactic acid bacteria, the need for magnesium was very low, in the range of 8.1 × 10 ⁻⁴ mM to 2.0 × 10 ⁻³ mM, only by incorporating it as an essential element. However, the present inventors have found that this magnesium content is 30 mM or more, particularly preferably 50 mM.
It was found for the first time that the inclusion of the above in the culture medium promotes the production of lactic acid and enhances the vitality of the starter. When the amount of magnesium exceeds 50 mM, the effect reaches a plateau, so even if more magnesium was added, there was no great difference in the effect. As the magnesium added to the culture medium, either an organic salt or an inorganic salt can be used, but an inorganic salt is particularly preferable. Further, examples of the inorganic salt include carbonate, hydrochloride, sulfate and the like. However, when the starter is used in foods, sulfates and hydrochlorides approved as food additives are desirable. The starter of the present invention is a raw material, skim milk powder or whey powder, calcium carbonate or calcium phosphate,
A magnesium salt and other additives are dissolved in a predetermined concentration and heat-sterilized to prepare a culture medium. If necessary, the prepared culture medium can be spray-dried to obtain a dry powder. In the cheese production, the thus-prepared culture medium is used in the same manner as an ordinary starter culture medium to prepare a starter, and this starter is added to the raw material milk and fermented to enable efficient cheese production.
In particular, the starter prepared with the culture medium according to the present invention has high vitality, and thus the time until collecting the curd can be shortened. Hereinafter, the present invention will be described in more detail by showing test examples and examples.

[0009]

[Test example]

(1) Lactic Acid Bacteria In this test example, the following strains were used as lactic acid bacteria and used as test strains. A commercially available starter was purchased, and lactic acid bacteria were separated from this starter. The isolated strain was identified according to a conventional method. The bacteria used in the test were L. lactis ssp.cre.
moris and below L. cremoris), Lactococcus lactis
Subspecies lactis (L.lactis ssp.lactis or less L.lactis), leuconostoc (Leuconostoc or less Le
u.). In the test of the starter, a mixed culture of these 3 strains was used, but if necessary, a single culture was used. When used alone, the culturing temperature condition for each lactic acid bacterium was 30 for L. cremoris and L. lactis.
℃, Leu. Was 25 ℃. 10% for commercial starters
After carrying out subculture for 5 or more generations with reduced skim milk (RSM) at a concentration, each lactic acid bacterium was separated, and the isolated bacterium was cultured using MRS medium (manufactured by Difco).

(2) Activity measurement In the present invention, the lactic acid degree was used as the activity of the starter. For vitality measurement, 10% reduced skim milk (R
(SM) was inoculated with 1% and cultured at 30 ° C. for 6 hours, and the lactic acid acidity was measured by the acidity titration method. Further, the ratio of the acidity of the test sample to the acidity of the culture, which was used as a reference for the relative activity, was calculated and expressed as relative activity (%). The pH and the number of growing lactic acid bacteria were also measured as a reference for vitality. The sterilization of 10% reduced skim milk to measure vitality is 100 ° C 3
The heating was performed for 0 minutes, and other sterilization was performed at 115 ° C. for 20 minutes.

(3) Measurement of vitality of starter prepared in a commercial medium A starter was prepared using a commercially available medium for lactic acid bacteria. The medium used was 10% reduced skim milk (RSM), M1.
7, MRS, LMB, GYP, Eliker medium. These culture media were prepared as instructed, and the above-mentioned three lactic acid bacteria strains were inoculated into each culture medium according to the above-mentioned vitality measurement method, followed by culturing to measure the vitality. The results are shown in Table 1 below.

[0012]

[Table 1] Vitality of starter prepared with lactic acid bacterium medium ────────────────────────────────── Medium vitality (lactic acid Acidity) Viable cell count (CFU / ml) ────────────────────────────────── 10% RSM 0.524 3.1 × 10 ⁸ M17 0.524 3.1 × 10 ⁸ MRS 0.450 7.2 × 10 ⁷ LMB 0.428 8.6 × 10 ⁷ GYP 0.448 9.6 × 10 ⁷ Eliker 0.395 6.6 × 10 ⁷ ────────────────── ─────────────────

When any medium is used as a starter, 1
It showed vitality equal to or less than 0% RSM.
One of the reasons why the commercially available medium showing the highest activity cannot exceed RSM is that it is not possible to suppress the decrease in pH due to lactic acid produced. Therefore, it has been attempted to add a neutralizing agent to the culture solution in the culture of lactic acid bacteria. This time, again, a study was conducted on a sparingly soluble salt that is added to the culture solution and has a pH-preventing effect.

(4) pH buffering action of lactic acid by sparingly soluble salts Calcium carbonate, magnesium carbonate, calcium citrate and magnesium phosphate were selected as sparingly soluble salts. 100 ml of a 1% aqueous suspension of these salts was prepared, and lactic acid having a concentration of 88.5% was added dropwise with stirring to measure pH change. The results are shown in Fig. 1. Optimal growth of lactic acid bacteria p
H is known to be in the range of 5.5 to 6.5,
From the results shown in FIG. 1, calcium carbonate and calcium phosphate were selected as the optimum salts for maintaining such a range.

The cause of the low activity in MRS is the decrease in pH. Therefore, a culture solution was prepared by adding the above-selected calcium carbonate or calcium phosphate to the MRS medium to a concentration of 1.5%, and from this culture solution, a starter was prepared in the same manner. 10 as a control
% RSM was used. The results are shown in Table 2.

[0016]

[Table 2] Effect of adding calcium carbonate and calcium phosphate to MRS medium ─────────────────────────────────── Medium Salt Vitality (lactic acidity) ────────────────────────────────── 10% RSM ── 0.56 MRS 1.5% ( Calcium carbonate) 0.56 MRS 1.5% (calcium phosphate) 0.58 ───────────────────────────────────

The addition of 1.5% calcium carbonate or calcium phosphate improved the vitality of MRS by about 20% over that shown in Table 1. This is because the added salt above
This is to adjust the pH to the optimum pH for lactic acid bacteria. Therefore, it has been found useful to add the above-mentioned sparingly soluble salt to the components of the culture medium for starters.

(5) Examination of nitrogen source and carbon source In the conventional starter preparation, 10% RSM has been empirically used. However, this concentration causes various problems as described above. Therefore, skim milk powder or whey powder was used as a nitrogen source and a carbon source, and the optimum concentration for growth of lactic acid bacteria was examined without causing coagulation. As a result, it was found that coagulation did not occur and growth of lactic acid bacteria was not affected by setting the skim milk powder and whey powder to 3.5% concentration. From the above results, the compositions shown in the following Table 3 were used in the following experiments as the basic composition of the culture medium for starters.

[0019]

[Table 3] Basic composition of culture medium for starter ─────────────────────────
─────Skim milk powder or whey powder 3.5% Yeast extract 0.5% Ammonium dihydrogen phosphate 1.5% Trisodium citrate 1.5% Calcium carbonate or phosphate 1.5% Calcium ─── ──────────────────────
─────

The effect of the combination of the culture medium and the salt having the above composition on the starter vitality was compared. 10% as a control
RSM was used. The results are shown in Table 4.

[0021]

[Table 4] Vitality of culture medium for starter ─────────────────────────────────── C source, N source Salt starter pH Vitality (lactic acidity) ────────────────────────────────── 10% RSM-4.6 0.60 Whey powder 3.5% Calcium phosphate 5.1 0.64 Whey powder 3.5% Calcium carbonate 6.1 0.64 Skim milk powder 3.5% Calcium phosphate 5.1 0.68 Skim milk powder 3.5% Calcium carbonate 5. 7 0.60 ────────────────────────────────────

As described above, 3.5 as a nitrogen source and a carbon source.
By using non-fat dry milk or whey powder with a concentration of 10%, it was possible to obtain an effect equal to or higher than that of the conventional 10% RSM. IP culture medium using calcium carbonate and whey powder
The culture medium using CM-W, calcium carbonate and skim milk powder is abbreviated as IPCM-S below.

(6) Addition Test of Magnesium for Vigor Enhancing Factors In order to further improve the effect of the above-mentioned culture medium, the factors for enhancing vitality were examined. When a soluble salt of magnesium was used as an inorganic salt in the examination of the above test, a phenomenon was observed in which the acid productivity per lactic acid bacterium increased. Therefore, magnesium sulfate and magnesium chloride were selected as the inorganic salts of magnesium and added to the above culture solution to confirm the effect. In each case, the culture medium was prepared, added before heat sterilization, sterilized, and inoculated with lactic acid bacteria. The results are shown in Table 5.

[0024]

[Table 5] Effect of magnesium salt as a vitality enhancer ─────────────────────────────────── Culture medium magnesium Salt pH pH after culturing (lactic acidity) ─────────────────────────────────── 10% RMS ── ─ 4.61 0.56 ICPM-S 5mM Magnesium Sulfate 5.18 0.52 ICPM-S 50mM Magnesium Sulfate 5.00 0.71 ICPM-S 5mM Magnesium Chloride 5.40 0.56 ICPM-S 50mM Magnesium Chloride 4.97 0.68 ICPM-S ──── 5.73 0.44 ────── ──────────────────────────── A remarkable activity-enhancing effect was confirmed by the addition of magnesium. It was also confirmed that the effect of this magnesium salt tends to be further enhanced by heat sterilization together with the culture medium.

[0025]

Example 1 In consideration of the above test example and use as a food, the following composition was determined as an example of a culture medium for a starter.

[0026]

[Table 6] ─────────────────────────
───Skim milk powder or whey powder 3.5% Yeast extract 0.5% Ammonium dihydrogen phosphate 1.5% Trisodium citrate 1.5% Calcium carbonate 1.5% Magnesium sulfate 50 mM ────── ────────────────────
────

Each component was dissolved in water, heat sterilized at 100 ° C. for 30 minutes, and cooled to room temperature to prepare a starter culture medium.

[0028]

Example 2 The culture medium for starter prepared in Example 1 (I
PCM-S) was used to prepare a starter for a single strain of lactic acid bacterium.
Lactic acid bacteria can be obtained from Fast-Slow from the above commercial starter.
They were separated using deferential agar and classified into Fast strain and Slow strain according to the magnitude of the lactic acid fermentation rate. These strains 1
The cells were cultured in 0% RSM, and 5 Fast strains and 2 Slow strains were used. This was transplanted as in the above test example, and the vitality and the number of lactic acid bacteria were measured. The vitality was expressed as the relative vitality when 10% RSM was used as the culture medium.

[0029]

[Table 7] ─────────────────────────────────── Strain 10% RSM IPCM-S ──── ─────────────────────────── Relative viability Viable count Relative viability Viable count (CFU / ml) (%) (CFU / ml) ─────────────────────────────────── Fast 1 100 5 × 10 ⁸ 140 1.0 × 10 ⁹ 2 100 6 × 10 ⁸ 131 9.0 × 10 ⁸ 3 100 3.8 × 10 ⁸ 148 9.2 × 10 ⁸ 4 100 5 × 10 ⁸ 120 9.0 × 10 ⁸ 5 100 4.2 × 10 ⁸ 129 1.0 × 10 ⁹ Slow 1 100 7 × 10 ⁷ 147 2.8 × 10 ⁸ 2 100 7.2 × 10 ⁷ 143 3.2 × 10 ⁸ ─────────────────────────────────── ─

The activity of the starter prepared from the culture medium of the present invention was high for each of the strains. The number of lactic acid bacteria is 2
It was confirmed that the culture medium of the present invention was suitable for the starter preparation, showing a value more than twice as high.

[0031]

Example 3 The starter culture medium for starter (IPCM-W) prepared in Example 1 was inoculated with a commercially available lactic acid bacterium starter to prepare a starter for cheese production, and its vitality was measured. As a control, a starter using 10% reduced skim milk as a culture medium was used. The vitality was determined by inoculating 10% reduced skim milk with a predetermined amount of starter and culturing at 30 ° C. for 6 hours, and measuring the lactic acid acidity. Table 8 shows the vitality of the control starter.
It was shown as a relative value as 0.

[0032]

[Table 8] ─────────────────────────────────── Culture medium Starter addition amount (%) Relative activity (%) ) ─────────────────────────────────── 10% RSM 1.0 100 IPCM-W 1.0 120 IPCM-W 0.5 91 IPCM-W 0.25 71 IPCM-W 0.01 59 59 ───────────────────────────── ──────

As is clear from Table 8, the starter prepared with the culture medium of the present invention has a strong vitality, and in producing cheese, the same effect was obtained with half the amount of the conventional starter.

[0034]

Example 4 The effects of the culture medium prepared in the example of the present invention and the commercially available culture medium for starter were compared. The components of the culture medium for the starter are unknown, but for the purpose of producing a starter for cheese, WIESBY (LABORATORIU
M WIESBY GmbH & Co. (KG, Germany)
There is a culture medium commercially available from This culture medium includes VIS as a starter preparation culture medium for cheese or fermented milk.
-There are two types of START 10 and 15. This commercial culture medium was purchased and compared with the starter culture medium of the present invention.
Inoculate with a commercially available lactic acid bacterium starter in the same manner as in Example 3,
A starter was prepared. Add 1.0% of this to the raw milk for cheese and keep it at 30 ° C overnight to obtain the lactic acidity and pH.
Was measured over time. Only the change in pH is shown in FIG. 2, but the starter prepared with the culture medium of the present invention was excellent in lactic acid fermentation satisfactorily and was clearly excellent in the decrease in pH.

[0035]

Example 5 Curds were prepared using the culture medium prepared in the examples of the present invention, and the effect on cheese production was confirmed. (1) Preparation of bulk starter 1% by weight of a mother culture prepared by culturing a commercially available cheese starter in a culture medium for mother culture prepared by dissolving sterilized skim milk powder in 10% concentration at 22 ° C. for 16 hours % To the above IPCM-W,
A bulk starter was prepared by culturing at 16 ° C for 16 hours (the present invention). As a control, skim milk powder was dissolved in 10% concentration,
Similarly, to a conventional starter culture medium prepared by sterilization, mother culture was added at a ratio of 1% by weight and fermented in the same manner to prepare a bulk starter (control).

(2) Card making 75 kg of 100 kg of raw material milk whose fat content was adjusted to 2.8%
The mixture was sterilized at 30 ° C. for 15 seconds, cooled to 30 ° C., put in a cheese vat, and 1 kg of the bulk starter was added. The lactic acid acidity was measured, and when it increased by 0.02%, 0.0038% of rennet dissolved in cold water was added. After forming the card, cutting and stirring were performed by a conventional method. One hour after the addition of rennet, 1/3 amount of whey was removed, and the following operation was performed in the same manner as in the normal Gouda cheese manufacturing method. When the lactic acid acidity of whey increased by 0.02%, whey was added. Exclusion was performed, cards were collected, and deposited and squeezed. The curd pressed for 25 minutes was cut, packed in a mold, pressed and shaped, and then immersed in cold water overnight, and transferred to an aging step as green cheese.

(3) Change in card formation state The card formation state was evaluated using the change in acidity during card formation as an index. As shown in FIG. 3, the increase in acidity was increased earlier in the present invention as compared with the control, and the time required for card making was reduced by about 1.5 hours. No difference was observed in the number of lactic acid bacteria in the green cheese obtained by the present invention and the control method.

Industrial Applicability According to the present invention, a novel culture medium for a lactic acid bacterium starter is provided. The starter prepared by the main culture medium has high vitality, the amount added during fermentation can be reduced, and the production cost can be reduced. Further, the fermentation time is shortened, and the fermented milk can be produced efficiently and stably. Also, the production cost of the starter can be reduced. Furthermore, when cheese is produced using the starter according to the present invention, the time required for card making can be shortened and the cheese production efficiency can be improved.

[Brief description of drawings]

FIG. 1 shows the pH change of a suspension of a sparingly soluble salt by lactic acid titration.

FIG. 2 shows the pH measurement results when a starter was produced using a commercial culture medium and a culture medium of the present invention.

FIG. 3 shows changes in lactic acid acidity when cheese is produced by the method of the present invention and a control method.

Claims (5)

[Claims] 1. A culture medium for the preparation of a lactic acid bacterium starter using milk as a nitrogen source and a carbon source, the pH of which is associated with lactic acid fermentation.
A culture medium for preparation of a lactic acid bacterium starter, which contains calcium carbonate or calcium phosphate in order to suppress the decrease in lactic acid, and which contains 50 mM or more of a soluble magnesium salt in order to promote the growth of lactic acid bacteria. 2. The milk is one or more substances selected from the group consisting of skim milk, whey, skim milk powder, and whey powder.
The culture medium for preparing the lactic acid bacterium starter described. 3. The culture medium for preparing a lactic acid bacterium starter according to claim 1, wherein the magnesium salt is magnesium sulfate or magnesium chloride. 4. The content of milk-derived solids in the culture medium is 3.5%.
The culture medium for preparing a lactic acid bacterium starter according to claim 1, which has a calcium carbonate or calcium phosphate content of 1.0% or more and a magnesium content of 50 mM or more. 5. A method for producing cheese, which comprises preparing a lactic acid bacterium starter using the culture medium according to claim 1 and preparing a cheese curd using the lactic acid bacterium starter.