JPH04341180A - Lactic acid bacterium containing highly unsaturated fatty acids and its production thereof - Google Patents

Abstract

PURPOSE:To effectively produce the subject lactic acid bacterium holding the original usefulness thereof, and further, facilitating the intake of a highly unsaturated fatty acid having a high nutritional and pharmacological value by culturing the lactic acid bacterium in a medium containing the highly unsaturated fatty acid. CONSTITUTION:Lactic acid bacterium is cultured in a medium containing a highly unsaturated fatty acid (e.g. linoleic acid or linolenic acid) or a fat or oil containing the unsaturated fatty acid (preferably an oil extracted from blue back fishes) to provide the objective lactic acid bacterium. The addition of the fat or oil to the medium in an amount of 0.002-0.1% is preferable, because the multiplication degree of the lactic acid bacterium and the intake degree of the highly unsaturated fatty acid are improved.

Description

[Detailed description of the invention]

0001

TECHNICAL FIELD The present invention relates to lactic acid bacteria containing highly unsaturated fatty acids and a method for producing the same. The lactic acid bacteria are useful in fields such as foods, cosmetics, agriculture, fisheries, and chemical products.

0002

[Prior art] Lactic acid bacteria have inherent pharmacological actions such as: ■ Proliferate in the intestine, produce substances that prevent the colonization of harmful bacteria, and occupy the colonization site of harmful bacteria. ■ Proliferate in the intestine. However, the bacterial components act as active substances, directly increasing the body's immunity, blocking the absorption of cholesterol, or improving the intestinal flora. It is widely used to suppress the production of bacterial intratoxins, enterotoxins, amines, ammonia, etc., but its bacterial fatty acid composition is mainly palmitic acid, palmitooleic acid, and oleic acid. Therefore, these fatty acids have poor nutritional and pharmacological value. Furthermore, it does not contain any highly unsaturated fatty acids.

On the other hand, highly unsaturated fatty acids such as linoleic acid and arachidonic acid are important as eicosanoid precursors, and are also known to have growth promoting and skin moisturizing effects. In addition, ω3 series highly unsaturated fatty acids such as α-linoleic acid, eicosapentaenoic acid, and docosahexaenoic acid are ω6
As an eicosanoid precursor associated with polyunsaturated fatty acids in the series, it has antithrombotic, antitumor, antiarteriosclerotic, and antiinflammatory effects, as well as lipid-lowering effects, and especially docosahexaenoic acid, which has effects such as suppressing vision loss and improving memory. Are known. All of these highly unsaturated fatty acids are essential fatty acids. Omega-6 series polyunsaturated fatty acids are found in terrestrial animal fats and oils, plant roots, etc., and can be consumed in sufficient amounts in the current diet, but omega-3 series polyunsaturated fatty acids, especially eicosapentaenoic acid and docosahexaenoic acid, is a fatty acid unique to marine organisms, and people of the generation who prefer meat eating tend to be deficient in it. A major reason why foods derived from marine organisms tend to be avoided is their aroma. Eicosapentaenoic acid and docosahexaenoic acid are also components of the aroma of seafood, but they are derived from aldehydes, which are oxidative decomposition products. Therefore, it is preferable that these highly unsaturated fatty acids can be added to foods in a form that is difficult to oxidize.

0004

[Problems to be Solved by the Invention] It is an object of the present invention to maintain the original usefulness of lactic acid bacteria and to improve the production of polyunsaturated fatty acids in order to facilitate the intake of nutritionally and pharmacologically valuable polyunsaturated fatty acids. An object of the present invention is to provide lactic acid bacteria containing the following.

[0005]

[Means for Solving the Problems] As a result of intensive research for the above purpose, the present inventors have discovered the following regarding the lipids of lactic acid bacteria cultured by adding fats and oils containing highly unsaturated fatty acids to the medium. I gained this knowledge. That is, (1) Even if the cells of lactic acid bacteria containing highly unsaturated fatty acids are washed with distilled water, a surfactant, and hydrochloric acid, the polyunsaturated fatty acids in the cells are as high as 30% or more of the total fatty acids; (2) The present invention The lactic acid bacteria used have a fast growth rate, and a large amount of bacterial cells can be obtained in a short period of time.

The present invention was completed based on the above findings, and is characterized by culturing lactic acid bacteria in a medium supplemented with highly unsaturated fatty acids or fats and oils containing them. Lactic acid bacteria containing fatty acids and a method for producing the same.

The highly unsaturated fatty acids used in the present invention include linoleic acid, γ-linoleic acid, arachidonic acid, and α-linoleic acid.
-Linoleic acid, eicosapentaenoic acid, docosahexaenoic acid and their salts, esters, triacylglycerol, diacylglycerol, monoacylglycerol, glycerophospholipid, glyceroglycolipid, sphingophospholipid, glycosphingolipid and the like.

[0008] The oils and fats containing these highly unsaturated fatty acids are preferably those in which the proportion of highly unsaturated fatty acids in the total fatty acids is 10% or more, and examples of such oils include sardines, mackerel, and horse mackerel. Preferred examples include oils and fats extracted from blue-backed fish such as tuna and bonito, oils and fats derived from the orbital fat of large marine fish such as tuna and bonito, oils and fats derived from microorganisms, and oils and fats derived from marine products extracted from squid liver.

[0009] The lactic acid bacteria that can be used in the present invention are not particularly limited in genus, species, or strain, but include, for example, Enterococcus, Lactobacillus, and Bihydrobacterium. These lactic acid bacteria can be easily obtained from public microorganism depositories.

[0010] In carrying out the present invention, lactic acid bacteria are cultured using a conventional method, for example, in a medium, and collected by centrifugation.
Wash and dry to obtain lactic acid bacteria powder. As the culture medium at this time, for example, a culture medium having the composition shown in Table 1 below can be prepared.

[0011]
Table 1 ──────────────
──────────────────
BHI (DIFCO)
3.7%
calf brain exudate
1.6%
beef heart infusion
2.0%
peptone
0.08
% Bactodextrose 0.02%
sodium chloride
0.04%
Sodium phosphate 0.02%
──────────────────────
─────────

pH 7.4

[0012] The amount of oil and fat to be added is 0.001% to 10%, preferably 0.002% to the medium.
% to 0.1% is desirable from the viewpoint of the growth of lactic acid bacteria and the uptake rate of polyunsaturated fatty acids. An emulsifier may also be added to the lactic acid bacteria culture medium.

[0013] When the bacterial cells cultured in such a medium are freeze-dried and then methyl esterified with hydrochloric acid and methanol by a conventional method, the fatty acid composition of all fatty acid derivatives ester-bonded in the bacterial cells can be analyzed by GLC. . Alternatively, the fatty acids constituting each lipid can be analyzed in the same manner after extracting wet or dry bacterial cells using an organic solvent and fractionating the lipids using silica gel TLC. By the above method, it was found that the fatty acids constituting the bacterial cells of the present invention are docosahexaenoic acid, eicosapentaenoic acid, arachidonic acid, linoleic acid, linoleic acid, oleic acid, palmitoleic acid, palmitic acid, myristic acid, etc. Ta.

[0014]

【Example】

Example 1 Enterococcus faecium
us faecium) IFO-3128 to BHI3.
7%, free fatty acids purified from tuna orbital fat [docosahexaenoic acid (95%), eicosapentaenoic acid (4%)]
)] Cells were cultured anaerobically in 1 L of a medium containing 0.05%, centrifuged and collected, and washed three times with distilled water, followed by 0.1% polyoxyethylene sorbitan monooleate.
The bacterial cells washed three times with Tween 80 (registered trademark) aqueous solution and three times with 6N hydrochloric acid were dried to obtain lactic acid bacteria powder.

[0015] Dry powder of lactic acid bacteria under each washing condition: 0.2
g was suspended in 8% hydrochloric acid methanol and reacted at 80° C. for 1 hour, and the resulting fatty acid methyl ester was analyzed by GLC to determine the content of highly unsaturated fatty acids.

[0016] In the bacterial cells that have been washed only three times with distilled water, the proportions of docosahexaenoic acid and eicosapentaenoic acid to the total fatty acids of the bacterial cells (content per 1 g of dry bacterial cells) are 87% (220 mg) and 3%, respectively. (8 mg).

Similarly, bacterial cells washed with a 0.1% polyoxyethylene sorbitan monooleate [Tween 80 (registered trademark)] aqueous solution had a concentration of 32% (14 mg) and 3% (
1.5 mg).

[0018] The bacterial cells washed with 6N hydrochloric acid were 36% (
19 mg) and 4% (1.9 mg).

Other fatty acid compositions are 0.1% myristic acid and 1% palmitic acid in the case of bacterial cells that have only been washed with distilled water.
8%, palmitoleic acid 0.6%, and vaccenic acid 4%. Similarly, bacterial cells washed with a 0.1% polyoxyethylene sorbitan monooleate [Tween 80 (registered trademark)] aqueous solution had a concentration of 1.2%, 12.8%, and 5%.
2% and 29.4%. Furthermore, the bacterial cells washed with 6N hydrochloric acid were 0.7%, 11.6%, 4.4%, and 29%.
It was 3%.

Example 2 Enterococcus faecalis
US faecalis) IFO-3181 to BHI3
．． 7%, free fatty acids purified from tuna orbital fat [docosahexaenoic acid (95%), eicosapentaenoic acid (4
%)] Medium 1 containing 0.01% monodocosahexanoylglycerol chemically synthesized from toglycerol
The cells were cultured aerobically in L, collected by centrifugation, and washed three times with distilled water. The cells washed three times and the cells washed three times with 6N hydrochloric acid were suspended in 5 mL of distilled water, and then extracted three times with 4 volumes of a chloroform/methanol (2:1) mixture.

Polar lipids were fractionated from the total fatty acids extracted from wet bacterial cells under each washing condition using silica gel TLC (developing solution: chloroform/acetone/methanol = 91:8:1), and 8% hydrochloric acid/methanol was added according to a conventional method. After reaction at 80° C. for 1 hour, the fatty acid methyl ester produced was analyzed by GLC to determine the content of highly unsaturated fatty acids. The proportions of docosahexaenoic acid and eicosapentaenoic acid (content per 1 g of dry bacterial cell) in the fatty acids constituting the polar lipids of the bacterial cell just washed with distilled water are 13% (26 mg) and 4%, respectively.
(8 mg).

Similarly, bacterial cells washed with 0.1% polyoxyethylene sorbitan monooleate [Tween 80 (registered trademark)] aqueous solution had a concentration of 12% (3.6 mg) and 4%.
(1.2 mg).

[0023] After washing with 6N hydrochloric acid, 9% (2
．． 7 mg) and 3% (0.9 mg).

[0024] Other fatty acid compositions are myristic acid 1.3% and palmitic acid 11% in the case of bacterial cells that have only been washed with distilled water.
．． 3%, palmitoleic acid 6.4%, vaccenic acid 52
%Met.

Similarly, bacterial cells washed with a 0.1% polyoxyethylene sorbitan monooleate [Tween 80 (registered trademark)] aqueous solution had a concentration of 1.1%, 11.2%, and 6.
1% and 54.6%. Furthermore, the bacterial cells washed with 6N hydrochloric acid were 1.4%, 11.7%, 6.7%, and 56%.
It was 3%.

[0026]

[Effects of the Invention] By culturing lactic acid bacteria in a medium supplemented with fats and oils containing highly unsaturated fatty acids, it has become possible to efficiently produce lactic acid bacteria containing highly unsaturated fatty acids in a short time.

Claims (2)

[Claims] [Claim 1] Lactic acid bacteria containing highly unsaturated fatty acids
2. A method for producing lactic acid bacteria containing highly unsaturated fatty acids, which comprises culturing the lactic acid bacteria in a medium supplemented with highly unsaturated fatty acids or fats and oils containing them.