JPH05252937A - Preparation of sour seed and nutrient medium for lactobacillus as a sour seed - Google Patents

Abstract

PURPOSE:To adjust the sour seed to the stabilized state, so as to cause no sensible change in the activity on subculture, thus facilitating the preparation of the culture medium and attaining good proliferation. CONSTITUTION:A lactobacillus for sour seed is inoculated in a medium to which cereal flour such as wheat flour, either acetic acid or sodium acetate or both of them are added so that they amount for 0.15 to 0.40wt.%, calculated as acetic acid content. When the subculture with a part of the seed is repeated, on every subculture, acetic acid or sodium acetate or both of them are added. Further, the nutrient culture medium for the lactobacillus as sour seed includes at least 3 nutrient components, namely, 1 to 2wt.%, based on the medium weight, of dried yeast essence, 5ppm to 100ppm of manganese, and 0.05 to 0.40wt.%, calculated as acetic acid, of acetic acid, sodium acetate or both of them.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for preparing sourdough seeds for baking and a nutrient medium for sourdough lactic acid bacteria for culturing sourdough lactic acid bacteria.

[0002]

2. Description of the Related Art In recent years, much attention has been paid to means for improving the quality and flavor of bread, and application of sourdough is considered as one means for that purpose. In the traditional sourdough bread method,
Lactic acid bacteria, yeasts, etc. naturally grown in each climatic climate have been densely grown in a medium using flour such as wheat flour, and this has been used as a fermenting seed for bread. The succession of the sourdough is usually carried out by a splicing operation in which grain flour such as wheat flour and water are further added based on a part of the sourdough to newly ferment.

Among these sourdoughs, there is known a production example of a sourdough which is used for culturing a specific lactic acid bacterium purely on the basis of a microbiological analysis (for example, Japanese Patent Publication No. 57-39734). Issue publication). In addition, some traditional sourdoughs have been subjected to treatments such as drying and freeze-drying, and preserved.
Distribution is made possible, which bakeries can use to make sour bread.

[0004]

[Problems to be Solved by the Invention] However, the traditional sourdough is capable of stable inheritance only in each specific area, and if it is replanted in different areas, the original sourdough It is known that activity is lost. This is due to the invasion of miscellaneous bacteria, which is different from the microorganisms that play a major role in the fermentation of each sourdough, and is due to the different microflora in the environment in different climates.

Further, the stability of the sourdough initially prepared can be improved by purely culturing the lactic acid bacterium and using it for seeding. However, when the subculture is repeated, the sourdough also has a similar cause. Problems tend to occur in which the nature of the seed differs from the beginning. Therefore, a stable method of inheriting sourdough that does not easily change its activity even after transplanting has been desired.

On the other hand, in the case of pure culture of lactic acid bacteria for sourdough, there is a complication that a special nutrient medium must be prepared depending on the bacterial species. Also, for example, several types of Lactobacillus san Francisco nutrient mediums used as lactic acid bacteria for sour seeds are known, but each has its own drawbacks. FYE (fresh yeast extract) is required in SDB medium (for example, published in Japanese Examined Patent Publication No. 58-58070), but this preparation work is time-consuming, and lactic acid bacteria may be produced depending on the type of raw yeast used as a raw material for FYE. Differences in growth may occur. In addition, it has been pointed out that there is a difference in the growth effect depending on the bacterial strain in a nutrient medium using L-cysteine as an alternative to FYE (for example, published in Japanese Examined Patent Publication No. 62-31907) (for example, Japanese Patent Laid-Open Publication No. 3-313). 172171 publication). Furthermore, although an example of culturing Lactobacillus San Francisco in MYP medium has been published (for example, published in Japanese Examined Patent Publication No. 3-58695), the amount of growth is not sufficient, and for the purpose of large-scale culture of bacterial cells, Not suitable. Therefore, there has been a demand for a nutrient medium in which the medium can be easily prepared and a good growth amount can be obtained.

The present invention has been made in view of the above points, and provides a stable method for preparing sourdough whose activity does not easily change even after splicing, and at the same time, a medium can be easily prepared and a good growth amount can be obtained. The purpose is to provide a nutrient medium that can be obtained.

[0008]

In order to achieve such an object, the method for preparing a sourdough of the present invention is, when preparing a sourdough by inoculating a flour medium such as flour with a lactic acid bacterium for sourdough, Either one or both of acetic acid and sodium acetate are added to the flour medium so that the initial acetic acid content is 0.15 to 0.40% by weight.

Further, one or both of acetic acid and sodium acetate are added to a flour medium such as wheat flour so that the acetic acid content is 0.15 to 0.40% by weight based on the flour medium, When sourdough lactic acid bacteria are inoculated to raise seeds of sourdough, and a part of this sourdough is subcultured and the culture is repeatedly passaged, one of acetic acid and sodium acetate is added for each subculture. One or both of them are added so that the acetic acid content is 0.15 to 0.40% by weight with respect to the initial flour medium.

When acetic acid or sodium acetate is added to the flour medium, the whole of the initial flour medium (flour, water, salt, etc., acetic acid and sodium acetate) is obtained in both sowing and subculture by subculture. 0.15 to 0.40% by weight, based on At the time of subculture, the values are based on the entire subculture medium (weight including acetic acid and sodium acetate) including a part of the previous sourdough.

Further, the nutrient medium of the present invention is a nutrient medium of lactic acid bacteria for sour seed used for pure culture of lactic acid bacteria for sour seed for inoculating sour seed, wherein 1 to 2 dry yeast extract is added per weight of the medium. % By weight, 5 pp manganese
m to 100 ppm, and either one or both of acetic acid and sodium acetate, or both of them, in an amount of 0.05 to 0.40% by weight as an acetic acid content, and at least three components are contained.

That is, the present invention provides for the preparation of sourdough
By adding either or both of acetic acid and sodium acetate to the flour medium in advance during preparation or subculture of sourdough, it is possible to stabilize the quality of sourdough during subculture. Also, in a pure culture of lactic acid bacteria for sourdough used for this, good growth can be obtained by using a nutrient medium in which one or both of acetic acid and sodium acetate are combined with other essential nutrients. Is.

[0013]

According to the method for preparing sourdough and the nutrient medium for lactic acid bacteria for sourdough of the present invention having such a constitution, either one or both of acetic acid and sodium acetate during the preparation or the subculturing are flour. By adding acetic acid to the medium in an amount of 0.15 to 0.40% by weight and adjusting the initial pH to around 5, the growth activity of lactic acid bacteria for sour seeds is improved and the contamination of various bacteria is prevented.
The three effects of improving pH buffering properties of sourdough are deliberately brought out simultaneously, thereby providing a stable method for preparation and subculture of sourdough. It also provides a nutrient medium for pure culture of lactic acid bacteria for sour seeds used for the sour seeds. The details will be described below.

(1) Improvement of activity of lactic acid bacteria constituting sourdough It has long been known that there are many Lactobacillus lactobacilli whose growth is stimulated by sodium acetate (Guirard, Snell and Willi).
ams: Arch. Biochem. 9, p361, 1
946) (Man, Rogosa and Sharp.
e: J. appl. Bact. 23 (1), p130,
1960). As a result of intensive investigations by collecting various lactic acid bacteria for sourdough, the present inventors fermented sourdough and can be used for the production of good sourbread. All lactic acid bacteria for sourdough promote growth by the presence of acetic acid. I found it. The effective acetic acid concentration varies depending on the bacterial species and the composition of the nutrient medium, but when the acetic acid content in the nutrient medium is generally within the range of 0.05 to 0.40% by weight, good results are obtained, The growth promoting effect can be confirmed by including acetic acid at any of these concentrations in the nutrient medium during pure culture of the seed lactic acid bacterium. Further, when acetic acid is added to sour seeds so as to have the above concentration, the growth of lactic acid bacteria for sour seeds can be promoted even in sour seeds. The addition of either one or both of acetic acid and sodium acetate to sourdough according to the present invention is, first of all, based on the above findings.

When culturing the bacterium, it is necessary that the pH is appropriate at the start of growth or fermentation, whether in pure culture or in sourdough. Either acetic acid or sodium acetate may be used to adjust the initial pH, or both may be used in combination. For example, when preparing sourdough, add acetic acid and sodium acetate in combination so that the acetic acid content does not exceed 0.4% and the initial pH is around 5. Thereby, a growth stimulating effect on lactic acid bacteria for sourdough can be obtained.

(2) Inhibition of various bacteria Next, the second advantage of adding one or both of acetic acid and sodium acetate to sourdough is suppression of various bacteria. Acetic acid has strong antibacterial activity and has been used as a food preservative since ancient times. Generally, sourdough is fermented from the initial pH of around 4.5 to 5.0, but acetic acid has a dissociation constant of pKa = 4.56 (25 ° C). The ratio of the non-dissociative molecules increases as the pH decreases, and the antibacterial activity also increases.

The addition of acetic acid to sourdough must be within a range that does not inhibit the activity of lactic acid bacteria that perform sourdough fermentation. According to the present invention, at least an inhibitory effect on sourdough bacteria can be expected, and the acetic acid-containing concentration by which lactic acid bacteria for sourdough can retain high growth activity is 0.15 to 0.40% by weight.
It turned out to be For example, specifically, acetic acid is added so as to have a concentration of 0.3% by weight with respect to the total amount of sourdough,
Also, by replacing part or all of acetic acid with sodium acetate and adjusting the initial pH of fermentation to around 5 with sodium hydroxide, etc., if necessary, the resistance to bacterial contamination can be significantly increased compared to the acetic acid-free section. Can be improved.

(3) Improvement of pH buffering property As a third advantage of adding one or both of acetic acid and sodium acetate to sourdough, the pH buffering property of sourdough can be improved. The improvement of the pH buffering property is particularly effective when the sourdough is to be produced with a liquid type in which the ratio of the amount of flour used to water is small. In the case of the liquid type, since the pH buffering property is lower than that of the sponge type or the like, the growth limit pH (3.
(8-4.0) is reached too early, and the fermentation ends with insufficient accumulation of metabolites due to fermentation. This is also the main cause of the lack of bread flavor and eventually the instability of sourdough performance. The use of acetic acid and sodium acetate as pH buffers for sour liquid species which also have the effects of (1) and (2) give good results. As an example of use, for example, by adding about 0.15 to 0.2% by weight of sodium acetate as acetic acid to a total of 100% sour liquid species containing 75% water and 25% wheat flour, a sponge species having substantially the same p
H-buffering is obtained.

[0019]

Embodiments of the present invention will now be described in detail with reference to the accompanying drawings.

(Example 1) Lactobacillus San Francisco: Lactobacillus for sourdough
Using the Sanfrancisco ATCC27651 and ATCC27653 strains, sodium acetate was added to the nutrient medium to confirm the growth promoting effect in pure culture. First, a known SDB liquid medium (FYE, that is, containing 0.5% of fresh yeast extract as a solid content, JP-B-58-5)
No. 8070), each bacterial cell was subcultured, collected at the end of logarithmic growth by centrifugation, washed, and resuspended in sterile distilled water. 2 × 10 ⁶ cells of these lactic acid bacteria suspensions
As shown in FIG. 1, each nutrient medium was inoculated so that the amount became / ml, and static culture was performed at 27 ° C., and the growth amount was measured with time. The growth amount was 600 nm OD (Optical) using each medium not inoculated with lactic acid bacteria as a control (reference).
Density) was measured by a general method. The results of the ATCC27651 strain are shown in FIG.

As shown in FIG. 2, it was confirmed that the growth of Lactobacillus san Francisco ATCC 27651 in the test medium c was faster than any of the basal medium, the test medium a and the test medium b, and the growth promoting effect of sodium acetate was confirmed. .. Similar results are obtained when the ATCC27653 strain is used.

Further, acetic acid was added to the above test medium b containing no sodium acetate in the range of 0.05 to 0.70% by weight to test the growth promoting effect on each of the above strains with an initial pH of 5.6. When acetic acid was added at a concentration of 0.05 to 0.40% by weight, growth of the control test medium b or more was observed. The same result can be obtained by adjusting the initial pH of the medium to 5.6 by adding sodium acetate instead of acetic acid as acetic acid so as to have the same concentration.

(Example 2) As an incidental matter derived from Example 1, fresh yeast extract has been said to be an essential component in the culture of Lactobacillus san Francisco, but it is commercially available instead of fresh yeast extract. It was found that the use of yeast extract and the combined use of manganese sulfate and sodium acetate can give growth equal to or higher than that when fresh yeast extract is used. The test medium c of Example 1 was modified to test medium d,
As shown in FIG. 3, other nutrient mediums (SDB medium, cysteine-containing section, MYP medium) are used in the same strain and method as
And growth was compared. The results with ATCC 27651 are shown in FIG.

As shown in FIG. 4, Lactobacillus san Francisco showed the fastest and good growth in the test medium d. The growth rate of the nutrient medium supplemented with cysteine instead of sodium acetate did not reach that of the test medium d. The MYP medium is not suitable as a medium for growing a large amount of cells since the amount of yeast extract tends to be insufficient. Similar results are obtained when using ATCC27653.

(Example 3) After confirming that Lactobacillus san Francisco promotes growth in the presence of acetic acid or sodium acetate in the nutrient medium of pure culture, the cultured cells were then used to prepare sour seeds. Was prepared. The strain is Lactobacillus san Francisco:
Lactobacillus sanfrancisc
o ATCC 27651 strain and yeast Candida mirelli capable of fermenting sourdough in symbiosis therewith:
Candida milleri ATCC 60590
It was used. Lactobacillus san Francisco was inoculated into the test medium d of Example 2 at 10 ⁷ cells / ml, and the mixture was shaken or stirred at 27 ° C. for 14 to 16 hours. Yeast Candida mirelli is YPG medium (containing glucose 2%, peptone 2%, yeast extract 1%) 10
One platinum loop was inoculated per 0 ml and shake-cultured at 27 ° C. for 18 to 24 hours. After collecting and washing these cells by centrifugation,
According to the formulation shown in FIG. 5, the flour medium was inoculated. Sodium acetate and acetic acid were added to the test section of the present invention so that the total amount of acetic acid exceeded 0.15%. This was designated as sourdough I, which was fermented at 27 ° C. for 8 hours with continuous stirring so that wheat flour would not settle, then stored in a constant temperature chamber at 12 ° C. for 16 hours, and after a total of 24 hours, the formulation shown in FIG. According to the instructions, transplanting was performed. Using this as sourdough II, fermentation, storage, and transplanting were repeated in the same manner. At this time, in the test section, sodium acetate is added so that the acetic acid content is 0.15% by weight or more for each subculture.

In this process, each time the sourdough was subcultured, sampling was performed from the sourdough immediately after substituting after completion of fermentation / preservation, and the number of yeasts was measured on a YM plate medium to determine the number of lactic acid bacteria as SDB (FYE solid content 0). 0.5% content) Using a plate medium, the number of bacteria was counted from colonies for those showing 10 ³ cells or more per 1 gram of sourdough, and the sourdough was centrifuged at 8000 rpm for 10 minutes, and the supernatant was collected to give 0.45 micron. After filtering with a deproteinization filter, the components were analyzed by high performance liquid chromatography. The results are shown in FIGS. 7 and 8.

From FIG. 7, the number of bacteria of Lactobacillus san Francisco in the test plot of the present invention is always 3 to 3 in the control plot at any stage after the subculture of the sour seed in the process of subculture of the sour seed. It increased five times, and in the control group, various bacteria A grew after sourdough IV, and sour odor also disappeared and the unpleasant flour odor became stronger, whereas in the test group, various bacteria contamination occurred. I didn't see any. The strain Lactobacillus san Francisco used in this test is S
White and raised colonies are formed on the DB plate medium,
The miscellaneous bacteria A form transparent and slightly flat colonies in the same medium, and also grow in YM plate medium for yeast to form colonies much smaller than the yeast Candida mireri, so that they can be easily distinguished. Bacteria A is the pH of an aqueous suspension of wheat flour.
A lactic acid bacterium that frequently grows regardless of the manufacturer of wheat flour when fermented for a long time under weak acidity around 5, and as a result of isolation and identification by the present inventors, the lactic acid bacterium is distributed in the plant kingdom and produces dextran from sucrose. Lactobacillus confusus: Lactobacillus confu
It is determined to be sus. This is a major cause of alteration of sourdough at the time of transplanting, especially when trying to produce sourdough especially in liquid type, but the addition of sodium acetate can suppress the growth of this bacterium.

Further, FIG. 8 shows that the sourdough of the test group of the present invention has a higher accumulation amount of the fermentation metabolites than the sourdough of the control group. The improved pH buffering property in the test section also has a favorable effect on the fermentation metabolism of lactic acid bacteria for sourdough.

(Example 4) S for measuring the number of lactic acid bacteria of Example 3
Bacteria A were separated from the DB plate medium and the growth response to acetic acid was confirmed by pure culture. Acetic acid was added stepwise using the test medium b of Example 1 as the basal medium to prepare 6N-HC.
The initial pH was adjusted to 5.0 with 1 or NaOH and the growth was measured in the same manner as in Example 1. Based on the growth amount of the acetic acid-free section after 24 hours, the ratio of the growth amount to this is shown in FIG. Lactobacillus San Francisco ATCC 27651 strain for sour seeds is 0.05-0.4
The growth was improved by the addition of 0% acetic acid, and the growth of the miscellaneous fungus A that invaded and propagated in sourdough was delayed by 0.15% or more of acetic acid.

[0030]

As described above, according to the method for preparing sourdough of the present invention, either one or both of acetic acid and sodium acetate are added to the sourdough during preparation of sourdough or at the time of subculture. As for acetic acid, 0.15
Since it is added so as to have a content of 0.40% by weight, the growth activity of lactic acid bacteria for sourdough seeds can be improved, contamination of various bacteria can be prevented, and the pH buffering property of sourdough seeds can be improved. Can be improved. Further, according to the nutrient medium of lactic acid bacteria for sourdough, good growth can be obtained by using a nutrient medium in which one or both of acetic acid and sodium acetate are combined with other essential nutrients. Therefore, the subculture of the sourdough is stabilized, and by using the sourdough while substituting it, it becomes possible to produce a sourdough bread that is lower in cost and more responsive than the conventional one.

[Brief description of drawings]

FIG. 1 is a diagram showing the structure of a test medium according to Example 1 of the present invention.

FIG. 2 is a graph showing the growth amount of Lactobacillus san Francisco over time, showing the growth promoting effect by the addition of sodium acetate.

FIG. 3 is a diagram showing the structure of a test medium according to Example 2 of the present invention.

FIG. 4 is a graph showing the growth amount of Lactobacillus san Francisco over time, showing a comparison in various media.

FIG. 5 is a diagram showing a test sourdough composition according to Example 3 of the present invention.

FIG. 6 is a diagram showing a test sourdough composition according to Example 3 of the present invention.

FIG. 7 is a diagram showing changes in the viable cell count in sourdough.

FIG. 8 is a diagram showing changes in components in sourdough.

FIG. 9 is a graph showing the growth amount of Lactobacillus san Francisco and various bacteria A after 24 hours, with an initial pH of 5.
0 shows that the acetic acid concentration was changed.

Claims (3)

[Claims] 1. When preparing a sourdough by inoculating a floury medium such as wheat flour with a lactic acid bacterium for sourdough, one or both of acetic acid and sodium acetate is added to the floury medium for the first time to contain acetic acid. A method for preparing sourdough which is added so as to have an amount of 0.15 to 0.40% by weight. 2. To a flour medium such as wheat flour, one or both of acetic acid and sodium acetate is added to the flour medium so that the acetic acid content becomes 0.15 to 0.40% by weight, When sourdough is prepared by inoculating lactic acid bacteria for sourdough and subcultured by using a part of this sourdough and subculturing repeatedly, one of acetic acid and sodium acetate is selected for each subculture. Alternatively, both of them are used as the initial acetic acid content of 0.15 to the flour medium.
A method for preparing sourdough, which is added so as to be 0.40% by weight. 3. A nutrient medium for sourdough lactic acid bacteria used for pure culture of sourdough lactic acid bacteria for inoculating sourdough, wherein 1 to 1 dry yeast extract per weight of the medium is used.
2% by weight, 5 ppm to 100 ppm of manganese, and one or both of acetic acid and sodium acetate as an acetic acid content of 0.05 to 0.40% by weight
A lactic acid bacterium nutrient medium for sourdough containing at least three components.