JPH09248179A - Production of iron-containing yeast - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain an iron-containing yeast useful for a food, effective for iron-deficiency anemia, having accumulated iron in high content without undergoing growth inhibition in the presence of an iron compound in a high concentration by culturing a yeast in a medium containing iron and magnesium in specific concentrations. SOLUTION: A high-RNA-containing yeast belonging to the genus Candida [e.g. Candida utilis CS-7,529 (FERM BP-1,656), etc.] is cultured in a medium to which 700-2,000ppm calculated as iron concentration of an iron compound (e.g. ferrous sulfate) and >=20ppm calculated as magnesium concentration of a magnesium compound (e.g. magnesium sulfate) are added, to give the objective iron-containing yeast which has >=1% iron content in a cell, hardly undergoes growth inhibition even in the presence of the iron compound in a high concentration, is effective for treating iron-deficiency anemia and is useful for a food, etc.

Description

Detailed Description of the Invention

[0001]

FIELD OF THE INVENTION The present invention relates to a yeast for health foods, which contains 1% or more of iron in the yeast cells by culturing edible yeast of the genus Candida in the presence of iron compounds. Manufacturing method.

[0002]

2. Description of the Related Art In the recent boom in health, interest in vitamins as nutrients as well as minerals and trace elements is increasing. Among these, iron is one of the minerals that are said to be in short supply along with calcium in Japanese. It is said that children and pregnant women in the growing stage are more likely to be suffering from iron deficiency anemia because their demands increase. Therefore, iron in a form that is easy to drink and easily absorbed is required.

Iron has already been positively added to processed foods, etc., and usually ferrous sulfate, ferrous fumarate, ferrous succinate, etc. are used. However, these compounds originally have an iron odor, and therefore the amount added must be limited, and there is a problem in terms of absorption into the body. Therefore, what has recently attracted attention is an attempt to supply yeast with iron in the form of organic iron to reduce the odor of iron or to improve the absorbability into the human body.

As a method for producing iron-containing yeast, a method for producing microbial cells containing a large amount of iron by culturing edible microorganisms in a medium containing iron compounds (Japanese Patent Publication No. 9835/1986),
Method for producing organic ferrous iron-containing blood donor composition by culturing yeast in sugar-containing nutrient medium in the presence of iron compound (JP-A-58
No. 101686 gazette), an iron compound is added to a medium containing a carbon source and a nitrogen source, and the microorganism is cultivated to contain iron at about 1% or less, and the concentration of magnesium in the medium is A method for producing microbial cells which needs to be 10 mg / l or less (Japanese Patent Laid-Open No. 62-1340).
No. 83) is known. Furthermore, a method for producing a ferric iron-containing yeast cell in which iron is accumulated by dispersing cells obtained by culturing yeast in an aqueous solution containing an organic compound and a divalent water-soluble iron compound (JP-A-5-176758) No. publication) is known.

[0005]

As described above, the conventional method for obtaining iron-containing cells is to obtain the iron-containing cells by culturing and growing the microorganism in a medium to which an iron compound has been added in advance. Alternatively, the microorganism is cultivated and proliferated in a medium to which an iron compound has not been added in advance, and then the bacterial cells are collected and the iron compound is added to the microorganism to be incorporated into the bacterial cell to obtain an iron-containing bacterial cell. In the former method, iron cannot be added at a high concentration because the growth is generally inhibited by iron. Further, the added iron compound is oxidized during a relatively short time by aeration and agitation during culturing, and in particular, an insoluble compound such as iron phosphate is added between the phosphate ion added as a usual essential medium component. In addition to inhibiting the uptake of iron by microorganisms, in the separation of bacterial cells by centrifugation after the completion of culture,
Since there is no significant difference in the sedimentation speed between the insoluble matter and the bacterial cells, it is difficult to separate them from the bacterial cells. The presence of this insoluble matter hinders iron odor, color, and absorbability. On the other hand, the latter method is considered to be a means for avoiding the growth inhibitory effect of iron itself on microorganisms, but since the compounds of phosphoric acid and iron increase depending on the concentration and contact time of both, iron is previously added. Even in the method of collecting iron from a microbial cell cultured in a medium that has not been added and then bringing it into contact with an iron compound to take up iron, in order to facilitate the uptake of iron, high concentration of phosphoric acid and iron Since the compound is used, the production and removal of iron phosphate and the like have become a problem.

[0006]

Means for Solving the Problems As a result of earnest research for solving the above problems, the inventors of the present invention have found that yeasts, particularly high RNA-containing yeasts belonging to the genus Candida, are treated with high concentrations of iron compounds and magnesium. Upon culturing in the presence of the compound, it was found that the iron content was high and growth inhibition did not occur, and the present invention was completed. That is, the present invention provides a method for producing yeast containing 1% or more, preferably 2% or more of iron.

Hereinafter, the present invention will be described in detail. Although any yeast may be used in the present invention, a high RNA-containing yeast belonging to the genus Candida is particularly preferable. The high RNA-containing yeast referred to in the present invention is, for example, Japanese Patent Publication No. 56-468.
As disclosed in Japanese Patent Publication No. 24, etc., it means a yeast having an ability to generate and accumulate RNA in an amount of 12% by weight or more based on the weight of bacterial cells.
da utilis) CS-7529 (FERM BP
-1656), Candida Uchiris
a utilis) CS-7550 (FERM P-3
341) etc. can be illustrated.

The present invention has an iron concentration of 700 p in the medium.
pm-2000ppm, 20p as magnesium concentration
It is carried out by adding pm or more. The iron compound used in the present invention may be an organic or inorganic substance as long as it has suitable water solubility. Specific examples of such a source include ferrous citrate, ferrous fumarate, ferrous lactate and the like. Besides, ferrous sulfate, ferrous chloride, etc., which are used as medium components in ordinary microbial culture, can of course be used. Although various iron compounds can be used as described above, ferrous sulfate is most preferable because it is inexpensive and has a standard for food addition in view of industrial scale production. The addition amount of these is 700 pp as the iron concentration.
It should be m to 2000 ppm. If the concentration is lower than 700ppm, the amount of iron accumulated will be less than 1%, while 2
If it exceeds 000 ppm, growth inhibition will begin to appear, and the accumulation efficiency of the added iron will decrease. Examples of magnesium used in the present invention include magnesium sulfate and magnesium chloride. The concentration of magnesium is 2
It is necessary to be 0 ppm or more, and if it is less than 0 ppm, the magnesium required for RNA synthesis is insufficient, which adversely affects the yield of saccharide cells and iron accumulation.

In the present invention, a high content of iron is accumulated in the cells by culturing at a relatively low temperature of 21 to 24 ° C. in a medium component having a restricted phosphoric acid concentration,
Without leaving the insoluble inorganic iron compound in the medium,
It is also possible to produce a high iron-containing yeast containing almost no low-molecular iron compounds in the cells. Examples of the phosphoric acid used include potassium phosphate and ammonium phosphate, and the phosphoric acid concentration is preferably about 0.03% to 0.5%.

As the medium composition of the strain, the carbon source is glucose, sucrose, acetic acid, which is usually used for culturing microorganisms.
Ethanol, molasses, sulfurous acid pulp waste liquid, etc. are used, and as the nitrogen source, urea, ammonia, ammonium sulfate,
Ammonium chloride nitrate or the like is used. As the phosphoric acid and potassium sources, potassium phosphate, ammonium phosphate, potassium chloride and the like are used. Other trace metals include zinc,
Inorganic salts such as copper, manganese and iron ions are effective. Further, if necessary, organic substances such as corn steep liquor, casein, yeast extract, and peptone may be added.

The cells used in the present invention have a culture temperature of 20.
It can grow in the range of ~ 33 ° C, but it is 21 for iron accumulation.
Relatively low temperatures of -24 ° C are preferred. The reason for this is that RNA content generally increases in proportion to the growth rate,
The present inventors speculate that at the same growth rate, it is said that the lower the culture temperature, the higher the RNA content. The culture pH is 3.5 to 7.5, preferably 4.0 to 6.0, and the culture time is usually 20 to 30 hours, varying depending on the concentration of the carbon source. In the present invention, when a high RNA-containing yeast is used, since it has a high phosphate uptake ability, it solubilizes precipitates such as iron phosphate and absorbs iron as well, so that it is cultured in a medium to which a high concentration of iron compound is added. Even if it does, almost no growth inhibition is observed. Also,
For example, as described in JP-A-62-134083, it is known that it is necessary to lower the magnesium concentration in the medium in order to obtain iron-rich microorganisms, but according to the present invention, By making the magnesium concentration higher than in the known example, yeast containing 1% or more, preferably 2% or more iron can be easily produced.

[0012]

EXAMPLES The present invention will be described below with reference to examples.
The bacterial cell concentration was measured by taking a fixed amount of the culture solution, washing the bacterial cells twice with a centrifuge, and removing a portion of the bacterial cell at 105 ° C.
It was calculated from the weight after drying overnight. The iron content incorporated in the cells was measured by ashing the cells by the wet decomposition method and then by the atomic absorption method. In the following Examples and Reference Examples, the cell concentration and the concentration of iron taken into the cell were measured by the same method.

Example 1 Yeast culture and iron accumulation 100 ml of YPD medium (glucose 2%, polypeptone 2%, yeast extract 1%) was dispensed into a 500 ml Erlenmeyer flask and sterilized at 121 ° C. for 15 minutes, and then candy. 1 platinum loop inoculation of Da Uchiris CS7529, 30 ℃
The mixture was shaken and cultured for 24 hours to give an inoculum. The whole amount of this was inoculated into a 30-liter fermenter. Glucose 6.5 as the medium
%, Monoammonium phosphate 0.26%, ammonium sulfate 0.15%, magnesium sulfate 0.09%, potassium chloride 0.2%, manganese sulfate 2 ppm, zinc sulfate 2 pp
m, and 0.4 ppm of copper sulfate were used for batch culture.
The culture conditions were 15 l of liquid in the tank, 30 ° C. of culture temperature, 15 l / min of aeration, 400 rpm of stirring, pH 4.0 (automatic control by addition of ammonia). As for iron, ferrous sulfate is 5000 ppm (1000 ppm as iron)
Was first added to the medium. The culture was terminated when glucose was completely consumed. Also, Candida
Uchiris CS7529 is a mutant strain containing a high concentration of RAN (Japanese Patent Publication No. 56-46824), and its parent strain was cultured under the same conditions. As a result, Table 1
Candida utilis CS75 inside the cells as shown in
It was found that 29 had a larger amount of iron accumulation. The accumulated iron content was 1.16%.

[0014]

[Table 1]

Example 2 Effect of Phosphoric Acid Concentration Candida utilis CS7529 seed culture was carried out in the same manner as in Example 1, and the same medium composition and conditions were used in a 30-liter fermentor except for the medium of Example 1 and ammonium phosphate. Culture was performed. The concentration of ammonium phosphate is 0.26%,
It was changed to 0.234%, 0.208%, and 0.182%. The results are shown in Table 2. From this result, the concentration of monoammonium phosphate that does not inhibit the growth was 0.182% (0.15% as phosphoric acid). Below this concentration, the growth was significantly inhibited and the culture was not completed. Therefore, it was possible to obtain a culture solution having an iron content of 1% or more and not producing an insoluble iron compound by carrying out batch culture in the range of a phosphoric acid concentration that does not inhibit growth when 1000 ppm of iron is added.

[0016]

[Table 2]

Example 3 Investigation of culture temperature Candida utilis CS7529 seed culture and 3
Batch culture was carried out in the same manner as in Example 1 in a 0 l fermentor. The culture was performed while changing the culture temperature in the culture conditions to 30 ° C and 24 ° C. As a result, it was found that culturing at a low temperature of 24 ° C. allowed the iron to accumulate in the microbial cells at a concentration about twice as high. In this case, it was necessary to raise the concentration of ammonium phosphate to a concentration at which growth was not controlled, as compared with the case of culturing at 30 ° C. That is, by increasing the ammonium phosphate concentration during culture at 30 ° C. by 20%, iron-containing bacterial cells could be obtained at the end of culture with almost no insoluble matter being produced. Since there was almost no change in RNA content and cell yield at both temperatures, it is considered that phosphate is essential for iron uptake.

[0018]

[Table 3]

Example 4 Addition amount of ferrous sulfate concentration and intracellular iron concentration At a culture temperature of 24 ° C., the concentration of ferrous sulfate was 5000.
A medium added by changing the concentration to 10,000 ppm was prepared and cultured in the same manner as in Example 1. Ammonium phosphate was used at a concentration that would not hinder the culture. As a result, Table 4
As shown in (1), when the ferrous sulfate concentration was 10000 ppm, the intracellular concentration of iron was as high as 4.32%.
The culturing time is 27 to 30 hours, and the addition concentration of ferrous sulfate is 5000 to 10000 ppm (iron concentration is 100
In the range of 0 to 2000 ppm), almost no growth inhibition was observed. 15 l of a culture broth obtained by adding 5000 ppm of ferrous sulfate to the culture was collected by centrifugation, suspended in the same amount of distilled water and centrifuged again to collect the cells. Distilled water was added to make a bacterial cell suspension of about 7%, and in order to reduce the viable cell count, the bacterial cells were kept at 80 ° C. for 5 minutes and then dried with a spray dryer. The drying conditions were an inlet temperature of 235 ± 5 ° C., an outlet temperature of 100 ± 5 ° C., and a liquid passing rate of 2 l / hour. As a result, 380 g of iron-containing cells having an iron content of 2.11% were obtained.

[0020]

[Table 4]

Reference Example 1 Properties of Iron Compounds Iron-containing yeast cells obtained by culturing at 24 ° C. in Example 3 in order to estimate the form of iron present in yeast cells. The dried product was suspended in distilled water at room temperature for 1 hour, and then iron in the bacterial cell portion and the supernatant was analyzed. Table 5 shows the results. It was found that iron content was extracted with water and only about 10% came out of the bacterial cells. Therefore, it is suggested that iron exists in association with high molecular weight proteins.

[0022]

[Table 5]

[0023]

As described above, according to the present invention, by culturing a bacterium that is not easily growth-inhibited by iron in a medium to which a large amount of iron has been added, edible yeast cells having a high iron content have been accumulated. Can be manufactured. A feature of such a bacterium is that RNA can be accumulated at a high concentration. Further, by adjusting the concentration of phosphoric acid, which is one of the essential medium components, it is possible to reduce the contamination of inorganic iron components such as iron phosphate that are usually generated during culture.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁶ Identification code Agency reference number FI Technical display location C12R 1:72)

Claims (4)

[Claims] 1. An iron concentration of 700 ppm to 2 in the medium.
A method for producing an iron-containing yeast, which comprises adding 000 ppm and a magnesium concentration of 20 ppm or more to make an iron content of 1% or more in the cells. 2. The yeast is Candid
The method according to claim 1, wherein the yeast is a high RNA-containing yeast belonging to the genus a). 3. The culture temperature is 21 to 24 ° C.
And the manufacturing method according to 2. 4. The phosphoric acid concentration in the medium is 0.03% to 0.5.
%, The manufacturing method according to claim 1.