KR100557006B1 - Phytoestrogenic isoflavone-enforced arrowroot products fermented by lactic acid bacteria and thereof producing method - Google Patents

Abstract

The present invention relates to a fermented lactic acid bacterium fermented with isoflavone, which is a plant female hormone substance, and a method for preparing the same, which has the effect of lactic acid bacteria, and further enhances the efficacy of phytoestrogens, and has a detoxifying effect of hepatotoxicity and an inhibitory effect against breast cancer. 칡 relates to fermented products. According to the present invention, the prevention and treatment of menopausal disorders of plant female hormone substances with precautions such as inhibition of harmful bacteria growth of lactobacillus fermentation, reduction of lactose intolerance, reduction of serum cholesterol, improvement of diarrhea and constipation, premenstrual syndrome It can be applied to the material of products with prevention and treatment, anti-cancer effect and vascular disease prevention effect and the material of whitening effect products by tyrosinase activity inhibition.

Lactobacillus, 칡, brown root, fermented product, female hormone, isoflavones, liver detoxification, breast cancer, whitening effect

Description

칡 lactic acid bacteria fermented product reinforced with isoflavone, a plant female hormone substance, and a method for producing the same {PHYTOESTROGENIC ISOFLAVONE-ENFORCED ARROWROOT PRODUCTS FERMENTED BY LACTIC ACID BACTERIA AND THEREOF PRODUCING METHOD}

The present invention relates to a fermented lactic acid bacterium fermented with isoflavone, which is a plant female hormone substance, and a method for preparing the same, and more specifically, to obtain a milled product by hydrolyzing water in the curd, and then amylase and glucoamlylase, Enzymatic digestion by addition of cellulase and pectinase enzymes results in enzymatic digestion, followed by fermentation by lactic acid bacteria such as Lactobacillus, Bifidobacteria, Streptococcus or Loconostock, followed by fermentation. The present invention relates to a method for recovering phytoestrogens by extracting fermented products with ethanol, and to fermented lactic acid bacteria fermented products having enhanced efficacy of lactic acid bacteria and phytoestrogens by such a method.

Conventional studies on phytoestrogen have revealed that flavonoids that act as phytoestrogens are present in the ovary readily accessible in Korea. The major isoflavones identified in this study include daidzein, daidzin, and puerarin, which are well known phytoestrogens (Anderson et al. Am. J. Clin. Nutr., Vol. 70, No. 3, page 464 1999: Knight et al., Gynecol., 87, 897, 1996).

Phytoestrogen is structurally similar to the female hormone steroidal estrogen, which binds to the estrogen receptor and acts as an agonist or antagonist against estrogen (Messina et al., J Nutr. 131, 3095S ~ 108S, 2001). ), Isoflavones come in four forms: aglycone, a sugar-dropped glycoside, or glycoside, acetylglucoside, or malonylglucoside. The shape of the flavones is known to vary.

These isoflavones are hydrolyzed by β-glucosidase in the intestine and are absorbed in the form of an active glycoside, that is, aglycone. In humans, it is known that bioconversion into analogous structural analogues by intestinal microorganisms. Phytoestrogen is classified as a complex of nonsteroidal structures, unlike estrogen, a female hormone used as a chemical hormone replacement therapy (HRT).

In contrast, isoflavones present in soybeans are mainly included in soybean embryos, and the conventionally known technique using the same is characterized in that isoflavones having estrogen action are disclosed in Korean Patent Publication No. 2002-060070 (Hepatocellular Enhancer). However, lactic acid bacteria fermentation method is not used.

In addition, in the production method of high-purity isoflavone aglycone by fermentation from soybean embryos (Korean Patent Publication No. 2002-032078), soybean embryos are fermented using Bacillus licheniformis rather than lactobacillus to acetone and ethanol. After the extraction by adsorption to the adsorption resin is separated, not only the strain is completely different from the present invention, but also the raw material using soybean embryos can be said to be different from the present invention.

In the method for producing aglycone isoflavones using microorganisms (Korean Patent Publication No. 2002-007553), an isoflavone glycoside solution is added to an adsorbed resin to convert microorganisms that secrete beta-glucosidase to a column to convert to aglycone. The application material and the like are also not clear, and a method of using a lactic acid bacterium fermentation product using 칡 as in the present invention is not disclosed.

In addition, a method for producing aglycone of isoflavones (Korean Patent Publication No. 2001-027341) uses an Aspergillus strain or a Trichoderma strain, and it is possible to use a strain of lactic acid bacteria as in the present invention. There is a difference, and they do not use 과 as in the present invention.

In another conventional technique for separating and recovering protein and isoflavones from plant materials (Korean Patent Publication No. 2002-103921), protein extracts are prepared and contacted with a polar ion exchange to separate isoflavones in the extract with an ion exchange resin. A technique for producing an isoflavone isoflavone (Korean Patent Publication No. 2001-091098) and a method for separating and purifying isoflavone from soybean embryos (Korean Patent Publication No. 2001-016220) are recovered by using an adsorption resin column. In the improved isoflavone extraction method (Korean Patent Publication No. 2001-089863), a technique using an adsorbent resin is disclosed, but all of them do not use a lactic acid bacteria fermentation product, and are mainly separated using a resin. Is doing.

In other words, lactic acid bacteria fermented products are not used as in the technique according to the present invention, and alcohol extraction, acid hydrolysis, and ion exchange resin methods are mainly used. In addition, most of the published technologies mainly target soybean embryos or soybeans, and there is no content of biotransformation of puerin present in the soybean to make Dyzedine. Moreover, fermented soybean products using useful strains such as lactic acid bacteria There was no study applying the.

As already known, lactic acid bacteria have been applied to various products since it has been found to be effective in promoting human health and preventing aging due to intestinal harmful bacteria.

Lactic acid bacteria are largely classified into Lactobacillus, Bifidobacterium, Staphylococcus, and Loconok stock strains, and the organic acids produced through heterozygous and homozygous fermentation are slightly different. In other words, the difference between Lactobacillus and Bifidocaterium is that Lactobacillus has homomorphic fermentation and dysmorphic foot, whereas Bifidobacterium has only heteromorphic fermentation.

In general, the physiological activity of lactic acid bacteria include inhibition of proliferation of harmful bacteria, alleviation of lactose intolerance, lowering of serum cholesterol, anticancer activity, immunity, diarrhea and constipation, and vitamin synthesis.

Techniques for incorporating the effects of such lactic acid bacteria into various foods have been developed, but most of these techniques are commonly used to make lactic acid bacteria beverages in which lactic acid bacteria are inoculated into milk and milk powder is added to increase lactic acid bacteria. In other words, conventional lactic acid bacteria products are limited to the foods developed and manufactured as lactic acid bacteria fermented food in addition to the limited indigenous Korean foods such as kimchi, so considering the dietary life of Koreans, it is not easy to consume lactic acid bacteria fermented foods. It is not easy.

In addition, foods such as yogurt have been developed in consideration of Korean taste, but since this is also difficult to see suitable for Korean tastes, the reality is that there is absolutely insufficient food suitable for Korean tastes. In the case of patents for producing lactic acid bacteria beverages or lactic acid using cereals as a conventionally developed technology for solving such problems, Lactic acid fermented beverages derived from vegetables and herbal medicines using Lactobacillus plantarum (Korea Patent Publication) 2000-0055832), rice snow drink (Application No. 1996-042486), method for producing lactic acid fermentation product using grain type rice (public number 92-16030), method for producing liquid lactic acid fermentation product using grain flour (public number 97-14592) , A method of producing lactic acid using a soybean-derived component (application number 10-1999-0017375) and the like.

However, these methods all consist of mixing milk or adding sugars such as glucose.

As described above, the conventional techniques for fermenting lactic acid bacteria, while lactic acid bacteria exist by fermentation using 칡 as in the present invention, mention of technology for 칡 fermentation at the same time strengthening the plant female hormone substance by bioconversion during fermentation is mentioned. Not.

An object of the present invention is to overcome the limitations of the conventional techniques as described above, and to provide a substance for enhancing the efficacy and phytoestrogens of lactic acid bacteria.

Another object of the present invention is to sterilize the pulverized product obtained from bovine to add various enzymes to obtain bovine enzymatically decomposed product having the best growth of lactic acid bacteria.

Yet another object of the present invention is to provide a method for further increasing the growth of lactic acid bacteria.

Another object of the present invention is to inoculate fermented lactobacillus, Bifidobacteria, Streptococcus, Loconokstock lactic acid bacteria in the enzyme hydrolyzate to obtain fermented lactic acid bacteria fermentation and soybean powder or powdered milk in 10% ~ It is to provide a method for bioconversion of the isoflavones present in the 칡 to the active phytoestrogen substance by inoculating and fermenting by lactic acid bacteria after 20% addition.

One feature of the present invention for achieving the above object is to hydrolyze the water to obtain a pulverized product, the preliminary warmed the pulverized product at 60 ℃ and then cooled by heating for 10 minutes at 100 ℃, amylase, gluco Enzyme digestion can be obtained by adding an enzyme of amylase, cellulase or pectinase enzyme, and enzymatic digestion is carried out, and the bacterium is heat-treated to sterilize while enzymatically digesting the lactobacillus, Bifidobacteria, streptococcus or It is incubated for 24 to 36 hours by inoculating 1% of any one lactic acid bacterium among Rokonosstock lactic acid bacteria.

In the case of amylase, the addition enzyme is added as 100 unit to 300 unit / g 칡, and in the case of glucoamylase, 2.50 unit to 5.00 unit / g 첨가 is added, and in the case of cellulose or pectinase, 1% Is preferably added.

Here, in order to increase the proliferation of lactic acid bacteria may be fermented by inoculating lactic acid bacteria after adding 10% to 20% of soybean or powdered milk as an added material to the enzyme digestion. The soybean or powdered milk is added to the inoculated ground lactic acid bacteria is preferably fermented up to 36 hours.

In addition, when inoculating bifidobacteria with lactic acid bacteria, it is preferable to add 0.05% ascorbic acid during fermentation, and the enzyme decomposition is preferably performed for 90 minutes.

In addition, the extraction of the phytoestrogenic substance is preferably extracted with ethanol from the fermented lactic acid bacteria. In addition, in order to increase the purity of the fermentation, the extract may be dissolved again in ethanol, or the precipitate generated by storing the ethanol extract at -5 ° C to -10 ° C may be recovered.

Through the above method can be obtained fermented lactic acid bacteria fermented isoflavone is a plant female hormone substance. The lactic acid bacteria fermented product thus obtained may be used as lactic acid bacteria, health food materials and products, menopausal prevention women food materials and products, beauty food materials and products, cosmetic materials and products, atopy materials and products or ointment materials and products. have.

First, briefly explaining the main points of the present invention, water is added to the water to obtain a pulverized product, and then heated and allowed to cool, followed by decomposition by addition of amylase, glucoamyl acetate, cellulose, pectinase enzyme, and the like. Heat treatment is used to sterilize the enzyme.

Lactobacillus, Bifidobacteria, Streptococcus, and Roconostock Lactic Acid Bacteria Cultivated in Media Are Inoculated into These Sterilized Enzymatic Hydrolysates or Millet Grinds Not Hydrolyzed by Enzymes.

At this time, in order to increase the proliferation of lactic acid bacteria may be inoculated with lactic acid bacteria grown in the medium by adding soybean powder or powdered milk to the enzymatically-dissolved milled product.

Extraction of phytoestrogen is a fermentation product prepared by fermenting with lactic acid bacteria up to 120 hours, preferably 36 hours through the same method as above to freeze at -20 ℃ ~ -60 ℃ to dry by lyophilizer to extract. At this time, in order to extract the flavonoids acting as phytoestrogens, ethanol of 5 to 10 times is added to the lyophilized sample dry weight and extracted for 12 hours while warming.

Here, in order to increase the purity of the extracted flavonoids, it is possible to increase the purity by recovering the precipitate generated by cold immersion, that is, the extract is dissolved again in ethanol, or by storing the ethanol extract at -5 ℃ ~ -10 ℃. In addition, in order to increase the extraction yield, ethanol extraction may be performed again using the extraction residue.

Example 1

PH pH change during fermentation of lactic acid bacteria by enzyme digestion

2 to 5 times of water is added to the water and pulverized with a colloid mill to obtain a pulverized product.Then, pre-warm at 60 ° C. and then heat at 100 ° C. for 10 minutes to cool the mixture. Amylase (amylase, 100 unit ~ 300 unit / g) Viii) and glucoamyl ace (glucoamlylase, 2.50 unit ~ 5.00 unit / g iii) was added to saccharification. Cellulase and pectinase enzymes were added in an amount of about 1% for enzymatic degradation.

These enzymatic digestions were performed for 90 minutes and then heat treated to sterilize the enzymes. Lactobacillus, Bifidobacteria, Streptococcus, and Roconostock Lactobacillus incubated in medium were inoculated on sterilized enzyme hydrolyzate or lysate which was not hydrolyzed by enzyme and incubated for 24 hours.

The results of investigating the pH are shown in Table 1.

As shown in Table 1, ascorbic acid was inoculated with 0.05% ascorbic acid after fermentation. The pH was 6.0 before the fermentation, but the pH was 0.3 ~ 0.6 lower than that without the hydrolysis with enzyme. Therefore, the enzyme treatment of 칡 increased the growth of lactic acid bacteria appeared to decrease the pH.

It is believed that various sugars, such as glucose, produced by the hydrolysis of starch present in 칡 by enzymes, contributed to the proliferation of lactic acid bacteria. The enzyme was best mixed with amylase and glucoamylase, and cellulose and pectinase were lower than this.

Different sources of carbon are used depending on the type of cells. Song et al. (Korean Journal of Microbiology, Vol. 30, No. 1, p. 63, 2002) show that Bifidobacterial SH2 grows well in lactose and adds glucose and maltose. In one case, he said he grew well. Trehalose (Ling et al., J. Ferment. Bioeng., Vol. 80, p. 204, 1995) is a storage carbohydrate and an energy storage source, a saccharide in nature that protects against physicochemical stress. Park et al. (Korean Journal of Food Science and Technology No. 28, No. 3, p. 451, 1996) showed that the addition of trehalose under the culture conditions of lactic acid bacteria with and without trehalose added B. breve. Said to grow well. In addition, Lee et al. (Korean Journal of Agricultural Chemistry, vol. 41, No. 7, pp. 527, 1998) shows that the saccharified liquor is a substrate suitable for bifidus in the process of producing saccharified liquor for fermentation of bifidus. It was minutes.

Therefore, as shown in Example 1, it can be seen that the saccharides generated by hydrolysis of the starch present in the pan by enzymes were used for propagation of lactic acid bacteria.

PH Change during Fermentation by Lactic Acid Bacteria with and without Enzyme Treatment    Enzyme lactobacillus Amylace Glucoamylases Cellulose Pectin Mixed enzyme No treatment L 4.6 4.7 4.9 4.9 4.4 4.9 Elby 4.7 4.6 4.9 4.9 4.5 4.8 Elsie 5.0 5.1 5.2 5.3 4.9 5.3 EL 5.0 5.0 5.3 5.2 5.0 5.3 Elka 4.6 4.7 4.9 4.8 4.4 4.9 baboon 4.5 4.7 4.9 4.9 4.6 5.0 Vilong 4.4 4.5 4.7 4.9 4.3 5.1 Biai 4.6 4.8 4.8 4.8 4.5 5.2 BP 5.0 5.2 5.3 5.3 4.9 5.4 Beatty 5.2 5.2 5.6 5.3 5.0 5.4 Esp 4.7 4.9 5.0 5.0 4.6 5.1 Estee 5.1 5.1 5.3 5.1 5.0 5.3 Rokoff 4.7 4.9 4.9 4.8 4.6 4.8 Locome 4.8 4.9 5.0 4.9 4.8 4.8

Lactobacillus abbreviation name: Lactobacillus acidophillus, L. bulgaricus, L. confusus, L. reuteri, L. casei, B. breve, B. longum, B. infantis, B. bifidum, B. thermophyllum, Str. faecalis, St. thermophillus, Leuconostoc plantarum, Locome ( Leuconstoc mesentroides)

Example 2

Number of Lactic Acid Bacteria by Fermentation Time

As in Example 1, lactic acid bacteria were inoculated into each of the enzyme-transferred (칡) and non-enzymatically-transformed (시켰다) to ferment for 36 hours.

At this time, the change in the number of lactic acid bacteria according to the fermentation time is shown in Table 2, in order to simplify the notation, the number of lactic acid bacteria × 10 ¹⁰ or more to ⑥, × 10 ⁹ or more to ⑤, × 10 ⁸ or more to ④, × 10 ⁷ or more is indicated by ③, × 10 ⁶ or less, ②, and × 10 ⁵ or less are ①.

In general, lactobacillus treated with amylase and glucoaryllace showed better growth than lactic acid treated with cellulose or pectinase, and lactic acid bacteria showed the best growth with the enzyme mixed with amylase and glucoamylase. In addition, although the number of lactic acid bacteria was higher in fermentation 36 hours than in 12 hours of fermentation, most of the lactic acid bacteria had a number of × 10 ⁸ ~ × 10 ^{6 It} was necessary to grow the lactic acid bacteria better.

In general, amylase among enzymes is contained in several germinating seeds, which act on amylose in starch molecules to randomly hydrolyze 1,4-bonds to produce dextrins, which continue to dextrin It is enzymatically decomposed into maltose or maltotriose.

Glucoamylases are known to produce glucose directly by acting on starch and acting on the 1,4-linkage, 1,6-linkage, and 1,3-linkage of amylose and amylopectin (Kim, Dong-Hoon, Food Chemistry, KKD, 1990) ). The fact that these sugars act on the proliferation of lactic acid bacteria was mentioned in Example 1.

Changes in Lactic Acid Bacteria during Fermentation of Lactic Acid Bacteria enzyme Amylace Glucoamylases Cellulose Pectin Mixed enzyme No treatment Fermentation time 12 36 12 36 12 36 12 36 12 36 12 36 L ③ ④ ③ ④ ② ③ ② ③ ③ ⑤ ② ③ Elby ③ ③ ③ ③ ② ③ ② ③ ③ ④ ② ③ Elsie ② ③ ② ③ ① ② ① ② ② ③ ① ② EL ② ③ ② ③ ① ② ② ② ② ③ ① ② Elka ③ ④ ③ ③ ② ② ② ② ③ ④ ① ③ baboon ③ ④ ③ ④ ③ ③ ③ ③ ③ ④ ② ③ Vilong ③ ③ ③ ③ ③ ③ ② ③ ③ ④ ② ③ Biai ② ③ ② ③ ② ② ② ② ③ ④ ① ② BP ③ ③ ③ ③ ④ ③ ④ ③ ④ ③ ① ② Beatty ② ③ ② ③ ② ③ ② ② ③ ③ ① ② Esp ③ ④ ③ ④ ③ ③ ③ ④ ③ ④ ② ③ Estee ③ ④ ③ ④ ③ ③ ② ③ ③ ④ ② ③ Rokoff ③ ④ ③ ④ ③ ③ ③ ③ ③ ④ ② ③ Locome ③ ④ ③ ④ ③ ③ ③ ③ ③ ④ ② ③

Acronyms of Lactic Acid Bacteria: Lactobacillus acidophillus, L. bulgaricus, L. confusus, L. reuteri, L. casei, B. breve, B.L. longum, B. infantis, B. longum, B. thermophyllum, Str. faecalis, St. thermophillus, Leuconostoc plantarum, Locome ( Leuconstoc mesentroides)

Example 3

Fermentation of Lactic Acid Bacteria with Soybean and Powdered Milk

As in Examples 1 and 2, lactic acid bacteria were inoculated and fermented by lactic acid bacteria in fermented or non-enzymatically treated cells, but more lactic acid bacteria were required to grow. For this purpose, soy and milk powder were used as additives.

That is, after the enzyme lysed by the method of Example 1 was added in a proportion of 10% to 20% in consideration of the water content of soybean powder or milk powder, 1% of the lactic acid bacteria grown in the medium was inoculated for up to 36 hours. Fermented.

At this time, the change in the number of lactic acid bacteria is shown in Table 3, in order to simplify the notation, the number of lactic acid bacteria × 10 ¹⁰ or more ⑥, × 10 ⁹ or more to ⑤, × 10 ⁸ or more to ④, × 10 ⁷ or more ③ is in the × 10 ⁶ or less ②, × 10 ⁵ or less was indicated, separated by ①.

As shown in Table 3, it was found that the fermentation of lactic acid bacteria by adding soybean and milk powder as a whole increased the number of lactic acid bacteria than that of Example 2, and the enzymatic digestion was good. Decomposition by amylase and glucoamylase among enzymes was better than treatment with cellulose or pectinase.

This indicates that the saccharides produced by enzymatic digestion, as mentioned in Examples 1 and 2, are used as carbon sources of lactic acid bacteria. Among the lactic acid bacteria, Lactobacillus and Bifidobacteria grew well in the milk-added mill, while Staphylococcus and Roconostok lactic acid bacteria grew well in the mill-added mill.

As long as it is widely known that lactic acid bacteria grow well in milk, it can be seen that proper protein and carbon sources are required for the growth of lactic acid bacteria. Cheng (J. Food Sci. 55, No. 4, No. 4, page 4, page 1178, 1990) used Streptococcus thermophilus and Lactobacillus casei to ferment lactic acid bacteria using soybean to make sogurt.

A study on the effect of heat treatment of soymilk on acid production of lactic acid bacteria and soybean yoghurt quality, Ko (Look. To produce an acid. Kanda et al. (Process. Biochem. 11, no.5, p. 23, 1976) inoculated Lactobacillus acidophilius by inoculating Lactobacillus acidophilius by heating soymilk made from soybean at 90 ° C, 100 ° C, and 121 ° C for 10 minutes or 20 minutes. I wanted to make it.

In the light of these studies, the addition of soybean or milk to crushed ground can lead to the proliferation of lactic acid bacteria.

Number of Lactic Acid Bacteria from Lactic Acid Bacteria Fermentation by Addition of Soybean or Milk Powder enzyme Amylace Glucoamylase Cellulose Pectin Mixed enzyme No treatment Fermentation time 12 36 12 36 12 36 12 36 12 36 12 36 bean Powdered milk bean Powdered milk bean Powdered milk bean Powdered milk bean Powdered milk bean Powdered milk bean Powdered milk bean Powdered milk bean Powdered milk bean Powdered milk bean Powdered milk bean Powdered milk L ④ ④ ⑥ ⑥ ④ ④ ⑥ ⑥ ④ ④ ⑤ ⑥ ④ ④ ⑤ ⑥ ④ ④ ⑥ ⑥ ④ ④ ⑤ ⑥ Elby ③ ⑥ ⑤ ⑥ ③ ④ ④ ⑥ ② ② ④ ⑤ ② ② ④ ⑤ ③ ④ ⑤ ⑥ ③ ④ ④ ⑤ Elsie ③ ③ ⑥ ⑥ ③ ③ ⑥ ⑥ ③ ③ ③ ④ ③ ④ ④ ⑥ ③ ③ ④ ⑤ ③ ③ ⑤ ⑤ EL ④ ④ ⑥ ⑥ ③ ③ ④ ④ ③ ④ ④ ④ ② ③ ④ ④ ④ ④ ⑥ ⑥ ③ ④ ④ ④ Elka ④ ④ ⑥ ⑥ ④ ④ ⑥ ⑥ ③ ③ ④ ⑤ ② ③ ④ ④ ④ ④ ⑥ ⑥ ③ ③ ⑤ ⑤ baboon ⑤ ⑤ ⑥ ⑥ ⑤ ④ ⑥ ⑥ ③ ④ ⑤ ⑤ ④ ④ ⑤ ⑥ ⑤ ③ ⑥ ⑥ ④ ④ ⑤ ⑥ Vilong ⑤ ⑤ ⑥ ⑥ ⑤ ④ ⑥ ⑥ ④ ④ ⑤ ⑤ ④ ④ ⑤ ⑤ ⑤ ④ ⑥ ⑥ ④ ④ ⑤ ⑥ Biai ③ ④ ④ ⑤ ③ ③ ④ ④ ③ ④ ④ ④ ③ ④ ④ ④ ④ ④ ⑥ ⑥ ③ ③ ④ ④ BP ④ ④ ⑥ ⑥ ④ ④ ⑥ ⑥ ④ ④ ⑤ ⑤ ④ ③ ⑤ ⑥ ④ ④ ⑥ ⑤ ④ ④ ⑤ ⑤ Beatty ③ ④ ⑤ ⑤ ③ ③ ⑤ ⑤ ③ ③ ③ ④ ③ ④ ④ ⑤ ④ ④ ⑤ ⑤ ③ ④ ④ ④ Esp ④ ③ ⑥ ⑥ ④ ③ ⑤ ④ ③ ② ④ ④ ③ ③ ④ ④ ④ ④ ⑤ ⑤ ④ ③ ⑤ ⑤ Estee ③ ③ ⑤ ⑤ ③ ② ⑤ ④ ③ ④ ⑤ ⑤ ③ ③ ④ ④ ③ ③ ⑤ ⑤ ③ ③ ⑤ ④ Rokoff ④ ③ ⑤ ⑤ ④ ③ ④ ④ ③ ② ④ ④ ③ ③ ④ ④ ④ ③ ⑤ ⑤ ③ ③ ④ ④ Locome ④ ③ ⑥ ⑤ ④ ③ ⑥ ⑥ ③ ② ④ ⑤ ④ ③ ⑤ ⑤ ④ ③ ⑥ ⑤ ④ ③ ⑤ ⑤

Acronyms of Lactic Acid Bacteria: Lactobacillus acidophillus, L. bulgaricus, L. confusus, L. reuteri, L. casei, B. breve, B.L. longum, B. infantis, B. longum, B. thermophyllum, Str. faecalis, St. thermophillus, Leuconostoc plantarum, Locome ( Leuconstoc mesentroides)

Test Example 1

Changes in Phytoestrogen Substances by Lactic Acid Bacteria Fermentation

Analysis of isoflavones acting as phytoestrogen was performed by HPLC of JASCO, Japan. The column used was OMC-based YMC AM303 (4.6 x 250mm), and the mobile phase was a solvent mixed with acetonitrile containing 0.1% acetic acid and secondary distilled water containing 0.1% acetic acid in a 30:70 ratio. Was used.

The flow rate was adjusted to 1.0 mL / min and the injection volume was 20 μl. The detector was a UV detector with a wavelength of 254 nm and sensitivity of 0.32. The standard for isoflavones is from Sigma Co, USA. It was provided by Murphy (Iowa State Unveristy) and Dr Kim (Kyung Hee university). Isoflavones were extracted at room temperature for 24 hours using 70% ethanol containing 0.1% acetic acid in lyophilized lactic acid bacteria.

Extraction concentration was adjusted to a concentration of about 120mg / 5mL. Supernatant obtained by centrifugation at 10,000 rpm after extraction was used as HPLC analysis sample. After glycoside hydrolysis, extraction was performed by adding 15 mL of 1N hydrochloric acid to 0.5 g of lyophilized lactic acid bacteria fermentation, and hydrolyzing for 90 minutes in a heating block at 120 ° C. to convert the isoflavone glycoside into aglycone. After hydrolysis, the mixture was cooled to room temperature, methanol was added to adjust to 50 mL, and the mixture was stirred to elute isoflavone. The supernatant obtained by centrifugation at 10,000 rpm was used as an HPLC analysis sample.

Examples 1, 2, and 3 by adding 10-20% of soybeans to the enzyme digestion using the mixed enzyme and fermented by lactic acid bacteria up to 120 hours, preferably 36 hours lactobacillus , Bifidobacteria, Staphylococcus, Loconok stock strain was selected and lyophilized at -20 ℃ ~ -60 ℃ to obtain a frozen product.

Table 4 shows the results of analyzing phytoestrogens acting as female hormones in the lyophilisate.

Before fermentation, 1370 mg / 100 g of dyzein, 821 mg / 100 g of dyedzine, and 5489 mg / 100 g of puerin were present in phytoestrogens.

In comparison, soybeans were found to be 5.3 mg / 100 g of dyed agent, 18.9 mg / 100 g of dydazine, 9.9 mg / 100 g of Genistein, and 42.3 mg / 100 g of Genistin before fermentation.

As the fermentation progressed, the content of puerarin present in the mud rapidly decreased, and dyed zein increased rapidly. Also, as the fermentation progressed, the content of dydzin and genistin decreased.

This phenomenon can be said that the lactic acid bacteria are fermented to the bovine, while puerin and dyed jine is bio-converted to dyed zein, zenithin and genistein. In other words, the content of puerarin in 전 before fermentation was 5489mg / 100g, but when fermented with lactic acid bacteria, the content was reduced to 1400-3209mg / 100g, whereas the Dyzedine contained in 칡 was 1370mg / 100g before fermentation. After fermentation, it increased from 2375mg / 100g to 3758mg / 100g to show that the bioconversion from puerin and dyedzine to dyedzein.

Kim et al. (Arch. Pharm. Res. 21, No. 1, p. 17, 1998) reported in their study on the metabolism of flavonoids by enteric microorganisms that enterobacteriaceae convert puerin and dydazine to dyedzein. And rutin in buckwheat and the like are also converted to quercetin.

(J. Microbiol. Biotechnol. Vol. 12, No. 1, p. 8, 2002) is Bifidobacterium sp. The beta-glucosidase present in int-57 completely hydrolyzed the flavonoid glycosides during fermentation of soymilk. They said that fermented soymilk for 18 hours converted hydrolyzed Dydzin and Genistin to Dyzedine and Genistin.

Isoflavonoids such as dydazine, pomononetin and genistin are reported to be converted and hydrolyzed by enterobacteriaceae in various animals (Griffiths et al. Biochem. J. 130 vol. 141 1972: Batterham et al. Aust. J. Agric. Res. , Vol. 22, p. 131, 1971, Barz et al., Phytochem., Vol. 9, p. 1745, 1970). In the conversion process, it can be said that the useful substances with female hormone action were converted into Dyzedein and Genistein.

However, most of the studies of the above researchers used the intestinal microorganisms of animals without the use of lactic acid bacteria is a big difference from the results of the present invention. In addition, in the method for producing aglycone of the soflavones (Korean Patent Publication No. 2001-027341), a strain using lactic acid bacteria in the present invention is used in the strain of the genus Aspergillus or the genus Trichoderma. There are many differences with the strain, and they do not use the 칡, in the present invention is a big feature that mainly used 칡. In addition, most of the known techniques do not mention that fuerin is bioconverted to make Dyzedine, and is mainly targeted to soybean embryos or soybeans.

Changes in Isoflavonoid Contents during Fermentation of Lactic Acid Bacteria (mg / 100g) Dyed Jane DIDJIN Genistein Jennysteen Puerin 전 (before fermentation) 1370 ± 12 821 ± 34 Not detected Not detected 5489 ± 120 Beans (Before Fermentation) 5.3 ± 0.5 18.9 ± 1.9 9.9 ± 1 42.3 ± 5 Not detected L 3190 ± 120 103 ± 12 48.8 ± 2 3.3 ± 0.2 2397 ± 88 Elka 2818 ± 173 120 ± 21 49.3 ± 4 2.0 ± 0.3 2750 ± 39 baboon 3190 ± 134 98 ± 9 48.5 ± 3 2.4 ± 0.1 1400 ± 28 Biai 3758 ± 151 132 ± 9 48.0 ± 2 4.0 ± 0.2 1800 ± 45 Vilong 3224 ± 176 153 ± 11 47.2 ± 4 3.4 ± 0.4 2320 ± 101 Esp 3379 ± 189 110 ± 8 46.0 ± 3 5.3 ± 0.5 2201 ± 120 Rokoff 2375 ± 167 115 ± 8 47.3 ± 2 4.9 ± 0.1 3209 ± 88

Acronyms of Lactic Acid Bacteria: Lactobacillus acidophillus, L. casei, B. breve, B. infantis, B. longum, Str. Faecalis, Rokoff (Leuconostoc plantarum)

Test Example 2

Detoxification effect of hepatotoxicity

The experimental animals were fed with 60 male milk rats for 14 days as a standard diet, and then divided male rats with an average body weight of 98.0 ± 2.5 g into six groups of 10 male eggs by the ovarian method to make ethanol 25% solution at the same time for 5 weeks. 2g per 1kg body weight was orally administered, ethanol non-feeding group (control) was fed the same calorie sugar solution. Water on a standard diet (20% casein, 50% corn starch, 15% sugar, 5% cellulose, 5% corn oil, 3.5% mixed minerals, 1% mixed vitamins, 0.3% mesionine, 0.2% choline chloride) Suspended fermented extract was added to prepare 5% instead of water.

After 5 weeks of breeding, anesthesia was opened, blood was collected from the abdominal aorta, and centrifuged. Serum was used as a sample. Serum aminotransferase activity was measured using kit (Eiken), and glutathione peroxidase activity was determined by Paglia et al., J. Lab. Clin. Med., Vol. 70, page 158, 1967), measured the reduction in oxidized NADPH at 340 nm for one minute per mg of protein. The activity of glutathione S-transferase was determined by the method of Habig et al. (Habig et al., J. Biol. Chem., Vol. 249, p. 130, 1974). The absorbance of GSH-DNCB conjugates Calculated using the coefficients. The organs were collected immediately after blood collection, washed with physiological saline, and weighed.

The weight of liver was lower than that of control and ethanol-treated groups, and the weight of liver increased by about twice as much as that of rats fed ethanol. 칡 Fermentation has been shown to inhibit alcoholic liver progression in rats by alcohol.

Serum aminotransferase is an enzyme (Huh et al., J. Nutrition., Vol. 105, page 1975) that reflects degeneration or necrosis of hepatocytes because tissue leaks into the blood and increases its activity. Liver damage during alcoholism is also considered a direct toxic effect of alcohol (Wardlaw et al., Prespectives in nutrition. P. 246, 1996).

In the present invention, when the fermented fermented product is reduced the activity of aminotransferase (aminotransferase) indicates that the damage of liver tissue due to ethanol administration was also reduced by the administration of fermented fermented product.

Glutathione S-transferase binds to the SH group of glutathione (glutathione), making it easy to excrete, and is known to have a function of protecting the living body from lipid peroxidation reactions (Burk et al., Biochem. Biophys Acta., Vol. 618, page 35, 1980). When rats were fed 36% of their total calories for 40 days with ethanol, glutathione S-transferase activity increased, which is an adaptive response to protect tissues from oxidative damage induced by ethanol. As described by Oh et al. (J. Nutr. 128, 758 pages, 1998), glutathione S-transferase activity was increased in the case of ingestion of fermented yeast.

Therefore, it is possible to suppress the oxidative damage caused by ethanol intake by ingesting fermented products.

Glutathione peroxidase protects cellular components from oxidative damage by simultaneously reducing hydrogen peroxide and lipid peroxide (Chow et al., J. Vit. Nutr. Res. 50, p. 364). 1980). Among enzymes that remove oxygen radicals, enzymes that use glutathione are known to have a large protective effect against tissue damage caused by alcohol intake (Cho et al., Korean Journal of Food and Nutrition Science, vol. 24, pp. 859, 1995). It is believed to increase glutathione release by increasing glutathione synthase activity by increasing glutathione peroxidase activity (Yoshida et al., J. Biochem. 96 vol. 1391, p. 1984). In the present invention, the activity of glutathione peroxidase in the ethanol- 칡 fermentation fed group was lower than that of the ethanol-administered group, indicating that administration of the fermented fermentation can suppress liver damage due to alcohol.

Hepatotoxic Effects of Fermented Fermented Rice in Ethanol-Fed Rats Weight of liver (g) Aminotransferase (K. unit / mL) Glutathione S-Transferase (nmole DNCB / mg Protein / min) Glutathione peroxidase (reduced NADPH nmole / mg / min) Control 1.65 ± 0.12 39.00 ± 1.22 19.01 ± 1.12 1.65 ± 0.12 ethanol 2.85 ± 0.22 50.22 ± 2.39 19.12 ± 1.38 1.72 ± 0.02 L 1.35 ± 0.23 33.77 ± 3.27 24.22 ± 0.29 1.64 ± 0.10 baboon 1.23 ± 0.12 30.33 ± 2.42 24.00 ± 2.36 1.62 ± 0.08 Esp 1.30 ± 0.19 32.22 ± 2.28 23.59 ± 3.23 1.69 ± 0.02 Rokoff 1.30 ± 0.17 31.00 ± 1.99 23.95 ± 2.98 1.70 ± 0.02

Acronyms of Lactic Acid Bacteria: Lactobacillus acidophillus, B. breve, B. infantis, Str. Faecalis, Locov (Leuconostoc plantarum)

Test Example 3

억제 Inhibitory Effect of Lactic Acid Bacteria Fermentation on Breast Cancer

Soybean-added fermentation prepared by adding 10-20% of soybean to the enzyme degrading product using the mixed enzyme by the method of Examples 1, 2, and 3 and fermenting it with fermented lactic acid bacteria up to 120 hours, preferably 36 hours Some of Lactobacillus, Bifidobacteria, Staphylococcus, and Loconok stock strains in water were frozen at -20 ℃ ~ -60 ℃, and dried by freeze-dryer 5% of the rats. Sixty females were fed for seven days, and then adapted to the experiment. The eggs were divided into six groups of ten by the ingot method and reared for 25 weeks.

After 25 weeks of breeding, anesthesia and laparotomy were performed to determine the total number of mammary gland cancer tumors in the mammary gland, and the tumor volume was calculated by 4 / 3πr ³ . DMBA, known as a breast cancer carcinogen, was dissolved in cooking oil and administered orally to 25 mg oral animals in a 50-day-old experimental animal.

After DMBA administration, abdominal palpation was performed to diagnose mammary cancer. The period until the first tumor appeared after administration of the carcinogen was recorded for each experimental animal and divided into tumor-bearing experimental animals for the total number of days per treatment to calculate the period until the first tumor appeared. In addition, the tumor occurrence rate was calculated by calculating the ratio of the tumor-bearing experimental animals to the total experimental animals on the basis of DMBA treatment age.

Feed intake of experimental animals was not significantly changed, about 15g ~ 17g / rat / day. As shown in Table 6, the addition of the fermented fermented food in the feed, it was effective in reducing the number of tumors in the experimental animals. In addition, the first tumor appearance after DMBA treatment, a carcinogen, was significantly delayed due to the addition of fermented products, and the rate of tumors was lowered until the end of the experiment. The tumor volume was also small in rats treated with fermented fermented rice, which suppressed the development of tumors. Therefore, Lactobacillus fermentation shows that it has the effect of suppressing breast cancer that may appear in menopausal women.

Inhibitory Effect of Lactobacillus and Lactic Acid Bacteria Fermentation before Fermentation on Breast Cancer Number of tumors Number of days the first tumor appears Tumor Prevalence (%) Tumor volume (mm ³ ) Control 5.2 ± 0.2 53.2 ± 1.8 56 ± 1.9 9933 ± 122 Before fermentation 4.9 ± 0.3 57.0 ± 0.2 52 ± 0.2 9020 ± 102 L 3.9 ± 0.3 62.7 ± 0.8 46 ± 2.1 8518 ± 108 baboon 3.8 ± 0.2 66.3 ± 2.1 42 ± 1.4 8182 ± 218 Esp 4.2 ± 0.1 63.7 ± 1.8 48 ± 1.2 8690 ± 171 Rokoff 4.5 ± 0.1 60.0 ± 1.5 49 ± 1.3 8709 ± 123

Acronyms of Lactic Acid Bacteria: Lactobacillus acidophillus, B. breve, B. infantis, Str. Facecalis, Lococon (Leuconostoc plantrum)

칡 Fermented products fermented with lactic acid bacteria in accordance with these embodiments has the effect of inhibiting breast cancer as the dietary estrogen substance that acts as a female hormone during the fermentation process, liver fermentation effect, and various functional food materials, lactic acid bacteria material, phytoestrogens material Can also be used.

Example 4

Recovery of Phytoestrogens from Fermented Soybean and Fermented Lactic Acid Bacteria by Ethanol Extraction

Examples 1, 2, and 3 of Lactobacillus, Bifidobacteria, Staphylococcus, and Loconos stock strains were used in the fermentation products produced by fermentation with lactic acid bacteria up to 120 hours, preferably 36 hours. Selected and frozen at -20 ℃ ~ -60 ℃ to extract the phytoestrogens acting as female hormone in the drying dried by lyophilizer.

The total flavonoid content in the extract was 50% methanol in 1 g of lactic acid bacteria fermented fermentation product using hand method (Hand Eun Shim, Exploration of total flavonoid content and physiological activity in vegetable food, Graduate Thesis, Ewha Womans University, 1999). 60 mL of the solution was added and refluxed at 80 ° C. for 1 hour. After cooling, the mixture was filtered with filter paper (Whatman No. 2) 100 mL of 50% methanol and used as a sample solution.

10 mL of diethylene glycol (diethylene glycol) and 1 mL of a sample solution were mixed in a test tube, and then, 1 mL of 1N NaOH was added thereto, mixed again, and the absorbance was measured at 420 nm after standing at 37 ° C. for 1 hour. The blank test was treated in the same manner using a 50% methanol solution instead of the sample solution. At this time, a calibration curve was prepared using a standard solution to determine the flavonoid content.

To extract flavonoids acting as phytoestrogens, ethanol (5-10 times) was added to the freeze-dried sample dry weight and extracted for 12 hours while warming. The ethanol extract was concentrated under reduced pressure to investigate the total flavonoid content of the concentrate.

As a result, ethanol extraction was able to recover more than 90% of the phytoestrogens present after fermentation. The total content was 7680mg / 100g in Example 4 before fermentation, but 6170.8mg / 100g recovery rate was 80.3% by simple extraction with ethanol. However, the fermented products recovered 87-92%. It was judged that this is because the water-soluble glycosides present in the shoot are converted into aglycone by the lactic acid bacteria fermentation.

In general, most of the flavonoids are insoluble in water when sugar is hydrolyzed, and they are small insoluble in organic solvents such as ethyl acetate (Kim Dong-hoon, Food Chemistry, p. 68, Insukdang, 1990). There are many technologies using isoflavones disclosed in Korea, but there is no method of fermenting lactic acid bacteria and using the fermented product or extracting from fermented product.

On the other hand, in order to increase the purity, the cold immersion method, that is, the extract is dissolved again in ethanol, or the ethanol extract is stored at -5 ℃ ~ -10 ℃ precipitates are generated, the recovery will be able to increase the purity, in order to increase the extraction yield The ethanol extraction may be performed again using the extraction residue.

Recovery of Phytoestrogens by Ethanol Extraction before and after Fermentation Lactobacillus Before fermentation L Elka baboon Biai Vilong Esp Rokoff Recovery rate (%) 80.3 ± 2.7 92.8 ± 3.3 88.7 ± 1.9 92.0 ± 1.6 91.0 ± 2.1 93.2 ± 2.5 90.8 ± 3.3 87.2 ± 1.9

Acronyms of Lactic Acid Bacteria: Lactobacillus acidophillus, L. casei, B. breve, B. infantis, B. longum, Str. Faecalis, Rokoff (Leuconostoc plantarum)

Test Example 4

Inhibitory Effect of Ethanol Extracts on Tyrosinase Activity in Fermented Fruits

Tyrosinase inhibitory activity was well mixed with 0.2 mL of 0.175M sodium phosphate, 0.2mL of 5mM DOPA solution, 0.5mL of 칡 and 칡 fermented extract of Example 4, and then left at 35 ° C. (tyrosinase) 0.1 mL of enzyme solution (110 unit / ml) was added thereto to react for 2 minutes, and then absorbance (S) was measured at 475 nm. After measuring the absorbance, 0.1 mL of distilled water was reacted for 2 minutes, and then the absorbance (B) was measured at 475 mn. After measuring absorbance (B), 0.5 mL of distilled water was added, mixed well, and left at 35 ° C., and then 0.1 mL of tyrosinase enzyme solution (110 unit / mL) was added thereto, followed by reaction for 2 minutes, and then absorbance (C) at 475 nm. It was measured again and the tyrosinase inhibitory activity effect was calculated as the following formula tyrosinase activity inhibitory effect (%) = [1- (AB / C)] × 100.

Tyrosinase is known as an enzyme involved in the production of melanin pigment, a black spot substance. Tyrosine synthesizes melanin pigments from tyrosine via dopa-quinone.

Accordingly, in the cosmetic manufacturing and pharmaceutical industries using living organisms as a raw material, the control of enzymatic oxidation of tyrosinase has been of interest. Tyrosinase, monophenol, dihydroxy-L-phenylalanine: oxygen oxidoreductase (EC 1.14.18.1), the only major enzyme in melanin biosynthesis, is a copper-containing enzyme that uses a wide range of phenolic compounds as substrates. The enzyme first goes through the Mason-Raper pathway using tyrosine as a substrate during the synthesis of melanin pigment, which causes dark spots in vivo.

Therefore, if tyrosinase is inhibited, the amount of melanin produced by melanocytes in the human body can be reduced, thereby achieving a whitening effect. Tyrosinease inhibitors include 4-hydroxyanisole, hydroxyquinone, 4-hexylresorcinol, troplpne, cinnamic acid, benzoic acid, koji acid, etc., which are used to treat hyper-pigmentation in humans. There is no report about this.

As shown in Table 8, the effect of inhibiting the tyrosinase activity of the ethanol extract of lactic acid bacteria fermentation is increased by more than 90% after fermentation, compared to 79.3% before fermentation, and can be used as a tyrosinase activity inhibitor. In light of various studies that melanin pigment, a skin deposition pigment, is produced, lactic acid bacteria fermented ethanol extract can be used as a whitening material for cosmetics and soap.

Tyrosinase Inhibitory Activity of Fermented Strains of Lactobacillus and Lactic Acid Bacteria Fermented product type Before fermentation L Elka baboon Biai Vilong Esp Rokoff Inhibitory effect (%) 79.3 ± 7.2 96.8 ± 4.3 93.7 ± 2.9 97.0 ± 2.9 93.0 ± 3.3 92.9 ± 5.5 90.8 ± 3.9 91.9 ± 3.3

Acronyms of Lactic Acid Bacteria: Lactobacillus acidophillus, L. casei, B. breve, B. infantis, B. longum, Str. Faecalis, Rokoff (Leuconostoc plantarum)

In general, melanin is produced in the basal layer, the lowest layer of the epidermis, and moves up with the metabolism of the skin. If the condition of the skin worsens, the melanin pigment remains and becomes blemishes such as blemishes and freckles.

Therefore, it can be said that a certain whitening effect can be obtained if it has a product function which suppresses the melanin pigment migration to an epidermis while suppressing tyrosinase. In Experimental Example 4 lactic acid bacteria 칡 fermentation shows that tyrosinase inhibitory effect can be used as a whitening material.

In the above embodiment, lactic acid bacteria fermented product is a product and material fortified with phytoestrogen, and also a hepatotoxic detoxifying material and product using these fermentations, a breast cancer suppressing material and product, a whitening material and product due to inhibition of tyrosinase activity It can be applied to.

As described above, the present invention by using the above-described technical configuration to make the lactic acid bacteria fermentation by using the enzyme digestion and 칡 at the same time by converting the isoflavones present in 로 into active plant female hormone (phytoestrogen) material The detoxification effect of hepatotoxicity, the inhibitory effect on breast cancer, and the whitening effect of tyrosinase activity inhibition can be made. In addition, it is possible to obtain a functional material by applying the phytoestrogens recovered from the 칡 lactic acid bacteria fermentation by ethanol extraction.

In addition, there is a whitening effect by inhibiting tyrosinase activity recovered from the lactic acid bacteria fermentation product by ethanol extraction.

In addition, lactic acid bacteria fermented product according to the present invention can be used as a lactic acid bacteria as well as a variety of health food material, women food material, beauty food material, atopy material, cosmetic material, ointment material.

What has been described above is only one embodiment according to the present invention, and the present invention is not limited to the above-described embodiment, and the present invention belongs without departing from the gist of the present invention as claimed in the following claims. Anyone skilled in the art will have the technical spirit of the present invention to the extent that various modifications can be made.

Claims (10)

Water was added to the water to obtain a pulverized product, and the pulverized product was pre-warmed at 60 ° C., then heated at 100 ° C. for 10 minutes, and allowed to cool. Then, the enzyme of any one of amylase, glucoamyl acetate, cellulose, or pectinase enzyme was used. To obtain enzyme lysate through enzymatic digestion by addition, the lysate was heat-treated to sterilize the enzyme and then sterilized with lactobacillus, Bifidobacteria, Streptococcus or Loconosstock lactic acid bacteria after 1% inoculation of lactic acid bacteria. A method for producing lactic acid bacterium fermented product, fortified with isoflavone, which is a plant female hormone substance, characterized by fermentation. The method according to claim 1, wherein the amylase is added at 100 units to 300 units / g ,, the glucoamylase is added at 2.50 unit to 5.00 unit / g ,, and for cellulose or pectinace The method for producing lactic acid bacterium fermented product, fortified with isoflavone, which is a plant female hormone substance, characterized in that 1% is added. According to claim 1, Isoflavone is a plant female hormone substance characterized in that the fermentation by lactic acid bacteria inoculated after adding 10% to 20% of soybean or powdered milk as an added material to the enzyme-decomposed to increase the proliferation of lactic acid bacteria 칡 lactic acid bacteria fermented product production method. The method of producing a fermented lactic acid bacterium fermented isoflavone, which is a plant female hormone substance, characterized in that when fermenting bifidobacteria, 0.05% ascorbic acid is added. The method according to any one of claims 1 to 4, wherein the enzymatic digestion is performed for 30 to 90 minutes. 6. The method for producing lactic acid bacteria fermented lactic acid bacteria fermented with isoflavone, which is a plant female hormone substance, characterized in that the lactic acid bacteria fermented product is extracted with ethanol to extract and recover phytoestrogens. The method of claim 6, wherein the extract is dissolved in ethanol to increase the purity of the fermentation, or the phytoestrogens, characterized in that by extracting and recovering the phytoestrogens generated by storing the ethanol extract at -5 ℃ ~ -10 ℃ Method for producing lactic acid bacteria fermented product enriched with isoflavone, a female hormone substance. Fermented lactic acid bacteria fermented with isoflavone, which is a plant female hormone substance, which is prepared by the method of claim 1. 9. The lactic acid bacteria fermented lactic acid bacteria fermented with isoflavones, which is a plant female hormone substance, characterized in that hepatotoxic detoxification, breast cancer inhibition and tyrosinase activating action. delete