KR20140047982A - Bacillus amyloliquefaciens 140n producing 1-deoxynojirimycin and method for producing fermented soybean having anti-diabetic activity using thereof - Google Patents

Abstract

The present invention can provide a bacillus amyloliquefaciens 140N strain producing 1-deoxynojirimycin (DNJ), and produce soybean-fermented products comprising 1-deoxynojirimycin having anti-diabetic, anti-cancer, and anti-virus effects using the same.

Description

[0001] Description [0002] Bacillus amyloliquefaciens 140N producing 1-deoxynojirimycin, and a method for producing fermented soybean having antidiabetic activity using the same, which comprises producing Bacillus amyloliquefaciens 140N producing 1-Deoxynojirimycin and a method for producing fermented soybean having anti-diabetic activity using thereof}

The present invention relates to a strain producing 1-deoxynojirimycin and a soybean fermentation product comprising 1-deoxynojirimycin produced by using the strain.

1-Deoxynojirimycin (DNJ), a multiple hydroxylated alkaloid, is a glucose analogue in which the oxygen atom of the pyranose ring is replaced by an NH group. DNJ has been shown to exhibit important biological activities such as antidiabetic, anticancer and antiviral activity and to inhibit α-glucosidase (Asano et al., 2000; Watson et al., 2001; Romaniouk et al., 2004) . These diverse potential activities have led to the development and production of DNJ and analogs in the functional food and pharmaceutical industries.

Various procedures have been developed for separating DNJ from mulberry roots (Asano et al., 2001) since the development of chemical methods to obtain DNJ by hydrogenation of noriglimycin (NJ) . However, when DNJ is extracted from mulberry leaves, the amount of extracted DNJ is small, the amount of DNJ is inconsistent depending on growing season and region, the production cost is high because long growing time is required, and similar alkaloid substances (Kimura et al., 2007; Vichasilp et al., 2009; Zhu et al., 2010).

Thus, studies have been conducted on the production of DNJ using fermentation by microorganisms including Bacillus and Streptomyces species. Although S. subrutilus and Streptomyces sp. SID9135 has been known to produce DNJ, but the fermentation time is long and it is inadequate for direct application in the food industry (Hardick et al., 1991; Paek et al., 1997). Accordingly, there have been several studies reporting the production of DNJ from B. subtilis DSM704 and B. subtilis MORI, focusing on Bacillus sp., Which is directly applicable to the food industry (Stein et al., 1984; Kim et al., 2011).

Although many studies have been reported on the isolation and identification of DNJs and their derivatives derived from microorganisms, there have been few studies on the biosynthesis of DNJ such as biosynthetic enzymes and pathways. So far, DNJ biosynthesis studies have been obtained from the results of isotopic labeling experiments, and there have been very few studies to isolate the genes involved in DNJ biosynthesis and to identify the enzymes involved. Recently, an anticipated operon consisting of amino-transferase (GabT1), phosphatase (Yktc1), and oxidoreductase (GutB1) was isolated from B. subtilis MORI 3K-85, which is a minimal biosynthetic gene cluster of DNJ (Kang et al., 2011).

Considering DNJ biosynthesis and its utility as an? -Glucosidase inhibitor derived from Bacillus sp. , The development of functional foods containing DNJ itself or containing DNJ itself may be of interest. The availability of DNJ is supported by previous reports that some Bacillus species ( B. subtilis B2 and B. subtilis MORI) producing DNJ were derived from traditional Chinese fermented beverage (okara) and Korean traditional fermented foods (soybean and Chungkookjang) (Zhu et al., 2008; Kim et al., 2011). However, little research has been done on the application of DNJ-producing Bacillus to functional foods.

A variety of compounds exhibiting inhibition of? -glucosidase have been chemically synthesized or extracted from natural products such as xanthone derivatives and salacinol. However, this approach has been associated with side effects such as hepatic disorder and low yield (Murai et al., 2002). Thus, the alpha -glucosidase inhibitor is S. hygroscopicus var. limoneus Derived validamycin (Iwasa et al., 1970) and Actinoplanes sp. Although the actinomycetes have been developed with a focus on actinomycetes as in the case of acarbose (Degwert et al., 1987), these actinomycetes are not suitable for application in the food industry even though they produce an? -Glucosidase inhibitor.

In the present invention, the inventors disclose a DNJ biosynthesis gene cluster derived from B. amyloliquefaciens irradiated from Korean traditional fermented foods based on PCR reaction. The inventors also provide optimal culture conditions for the best DNJ production. The inventors also tried to apply B. amyloliquefaciens , which produces DNJ, to soybean fermented foods having enhanced? -Glucosidase inhibitory activity.

According to one embodiment of the present invention, in order to produce DNJ using B. amyloliquefaciens and functional fermented food using the same, 90 kinds of microorganisms derived from traditional foods were analyzed for DNJ biosynthesis gene group and UPLC-ESI-Q -TOF-MS and DNJ biosynthesis B. amyloliquefaciens was investigated by measuring α-glucosidase inhibitory activity.

In order to isolate the DNJ biosynthesis gene group, the internal gene of yktc1, which is one of the DNJ biosynthesis genes based on the conventional studies, was used as a probe. The gabT1 , yktc1 and gutB1 genes were separated by PCR screening, And the nucleotide sequence of one DNJ biosynthesis gene group was analyzed.

Optimization of carbon source and nitrogen source during the cultivation of B. amyloliquefaciens 140N was performed to optimize the inhibitory activity of? -glucosidase.

In order to develop α-glucosidase inhibitory activity and functional food containing DNJ, soybean fermented products were prepared using B. amyloliquefaciens 140N, and α-glucosidase inhibitory activity was determined by direct extraction of soybean fermented product .

According to an embodiment of the present invention, a method of culturing a cell line in a culture medium containing at least one carbon source selected from the group consisting of water-soluble starch, maltose, lactose, sucrose and glucose, At least one nitrogen source selected from the group consisting of casamino acid, peptone, yeast extract, tryptone, and whey powder.

According to one embodiment of the present invention, there is provided a fermented food comprising 1- deoxynojirimycin fermented with B. amyloliquefaciens 140N.

According to one embodiment of the present invention, there is provided a method for producing a fermented food comprising 1- deoxynojirimycin , comprising fermenting a fermented object with B. amyloliquefaciens , and in an embodiment of the present invention According to an embodiment of the present invention, the fermentation is performed at 30 to 42 ° C.

According to one embodiment of the present invention, a 1-deoxynojirimycin producing strain of B. amyloliquefaciens 140N (KCCM11286P) is provided.

As used herein, the term "soybean fermented food" means a food prepared by using the fermentation action of a microorganism having soybean as a raw material. Examples of soybean fermented foods include soybean paste, hot pepper paste, And soy sauce.

The strain producing 1-deoxynojirimycin according to the present invention produces a large amount of DNJ as compared with the conventional one, and thus the antidiabetic activity having a high DNJ content and? -Glucosidase inhibitory activity is reinforced Can be used to produce soybean fermented products having high functionality.

1 is a phylogenetic diagram prepared by analyzing a 16S rRNA gene sequence (1a) of a 1-deoxynojirimycin producing strain and an entire amino acid sequence (1b) of a DNJ biosynthesis gene group according to an embodiment of the present invention.
FIG. 2 is a result of analyzing the entire amino acid sequence of the DNJ biosynthesis gene group of strains according to an embodiment of the present invention.
FIG. 3 is a result of UPLC-ESI-Q-TOF-MS analysis of the culture supernatant of B. amyloliquefaciens strain 140N according to an embodiment of the present invention and the supernatant of soybean fermentation product prepared using the strain.
Figure 4 is a block diagram of an embodiment of the present invention. glucosidase inhibition of amyloliquefaciens 140N according to the carbon source.
Figure 5 is a block diagram of an embodiment of the present invention glucosidase inhibition analysis of amyloliquefaciens 140N according to the nitrogen source.
Figure 6 is a B. amyloliquefaciens 140N in accordance with one embodiment of the invention a 2% soluble starch, 1% tryptone, 0.05% KH ₂ PO _4, and 0.05% (NH ₂₎ 5L fermentor in a medium containing ₄ SO ₄ . The results are shown in FIG. Square: Gun cell weight, Source: DNJ yield

Hereinafter, the components and technical features of the present invention will be described in more detail with reference to the following examples. However, the following examples are intended to illustrate the contents of the present invention and are not intended to limit the scope of the invention.

Various embodiments described herein are described with reference to the drawings. In the following description, for purposes of complete understanding of the present invention, various specific details are set forth, such as specific forms, compositions, and processes, and the like. However, certain embodiments may be practiced without one or more of these specific details, or with other known methods and forms. In other instances, well-known processes and techniques of manufacture are not described in any detail, in order not to unnecessarily obscure the present invention. Reference throughout this specification to "one embodiment" or "embodiment" means that a particular feature, form, composition, or characteristic described in connection with the embodiment is included in at least one embodiment of the invention. Accordingly, the appearances of the phrase " in one embodiment "or" the embodiment "in various places throughout this specification are not necessarily indicative of the same embodiment of the present invention. In addition, a particular feature, form, composition, or characteristic may be combined in any suitable manner in one or more embodiments.

Example  1. Experimental Method

1.1. Bacterial strains and general procedures

Approximately 90 bacteria from Korean traditional fermented foods were used as genetic resources for PCR screening and cultured in media such as TSB and MRS. Escherichia for gene cloning coli DH5? were cultured in LB medium at 37 占 폚. If necessary, ampicillin (100 μg / ml), 5-bromo-4-chloro-3-indolyl-β-D-galactopyranoside and IPTG (isopropyl-β-D-thiogalactopyranoside) were used. PGEM-T Easy Vector Systems Kit (Promega, USA) was used for cloning and sequencing.

1.2 DNJ  Biosynthesis Gene group  detach

In order to isolate the DNJ biosynthesis gene group, primers (MJS-1: 5'-GGWCTTGAAATGTCDTGTATYTCAATGTCDTGTATYTCBATHGCD-3 'and MJS-2: 5'-AAABACRCTKGCVGCWATATCRTAWGG-3') prepared on the basis of the internal base sequence of the yktc1 gene, PCR amplification was performed. In order to identify the screened strains, the 16S rRNA gene was PCR amplified using 518F (5'-CCAGCAGCCGCGGTAATACG-3 ') and 800R (5'-TACCAGGGTATCTAATCC-3') primers. The following three primer sets were constructed to identify the DNJ biosynthesis gene family containing the entire ORF of GabT1, Yktc1 and GutB1; (5'-GTGAGAGACTACTATCATYGRGCTTGGA-3 ') and MJS-20 (5'-TCACTTGATTTCCTCCAATAGCTTGCG-3' TTAGGAGTCCAGACCAACGCCTTCATA-3 '), GutB1: MJS-21 (5'-ATGAAGGCGTTGGTCTGGACTCCTAAT-3') and MJS-22 (5'-TTATAAAAGTTYCGGATCAGACACRAG-3 '). The PCR was carried out at 94 ° C for 5 minutes, followed by 35 cycles of 94 ° C for 30 seconds, 55 ° C for 1 minute and 72 ° C for 1 minute, and finally at 72 ° C for 10 minutes.

1.3 Establishment of optimal culture conditions

B. amyloliquefaciens 140N seedlings were cultured in 37 ° C TSB medium for 24 hours. A culture inoculum of 1% glucose, 1% yeast extract, 0.05% KH ₂ PO _4, and 0.05% (NH ₂₎ were inoculated with 1% (v / v) in the culture medium (100 ml) containing ₄ SO ₄ 37 Lt; 0 > C for 48 hours. maltose, lactose and sucrose as a carbon source and malt extract and beef extract as a nitrogen source in order to select a carbon source and a nitrogen source for optimization of? -glucosidase inhibitory activity, , Soytone, casamino acid, peptone, yeast extract, tryptone and whey powder. Using optimized media, the amount of DNJ production over time of B. amyloliquefaciens 140N was observed using a 5L fermenter. 1% (v / v) of the seed culture was inoculated in 3 L of medium (2% water-soluble starch, 1% tryptone, 0.05% KH ₂ PO ₄ and 0.05% (NH ₂ ) ₄ SO ₄ ) Lt; RTI ID = 0.0 > 37 C < / RTI > for 48 hours.

1.4 Analysis Method

To measure the? -glucosidase inhibitory activity, the culture was heated for 10 minutes, centrifuged at 10,000 rpm for 10 minutes, and the supernatant was recovered. 147 μl of the supernatant was divided into each well of a microplate reader (Thermo Electron Corporation, USA), and then 30 μl of 0.1 M phosphate buffer (pH 7.2), 30 μl of 1.42 mM p -nitrophenyl α-D -Glucopyranoside (PNPG, Sigma-Aldrich, USA) and 5 [mu] l of 1.8 U / ml [alpha] -glucosidase. After 10 minutes of reaction at 37 ° C, 100 μl of 0.5 M Na ₂ CO ₃ was added to terminate the reaction, and the absorbance was measured at 405 nm. The inhibitory activity was calculated through the following equation;

α-glucosidase inhibition (%) = [1- (sample A ₄₀₅ - blank A ₄₀₅ ) / control A ₄₀₅ ] × 100

Here, the blank represents distilled water instead of? -Glucosidase, and the control group represents the case where a supernatant sample is used instead of? -Glucosidase.

(UPLC-ESI-Q-TOF-MS) was analyzed with an Agilent 1290 Infinity UPLC system and a hydrophilic chromatography (HILIC) column (ZORBAX HILIC Plus, 2.1 x 100 mm, Agilent Technologies, USA). Solvent A (water containing 5 mM ammonium acetate) and solvent B (acetonitrile with 0.1% formic acid) were injected at a flow rate of 0.3 ml / min for 0 min (90% B) , 0.1 minute (90% B), 9.9 minutes (60% B), 1 minute (100% B), 2 minutes (100% B), 1 minute Concentration gradient. The eluate was detected using an Agilent 6250 Q-TOF mass spectrometer (Agilent Technologies, USA) at a gas (N ₂ ) flow rate (8 L / min) and gas temperature (325 ° C.) and analyzed using the Agilent MassHunter data acquisition program Respectively.

1.5 B. amyloliquefaciens  Using 140N Soybean fermentation product  Produce

200 g of beans were placed in water overnight at room temperature, and then heated at 114 ° C for 50 minutes. The cultured soybeans were inoculated with 3 ml of B. amyloliquefaciens 140N or B. licheniformis 320 cultured in TSB medium for 3 days at 37 ° C and 70% relative humidity. After fermentation, distilled water was added to 10 g of soybean fermentation product and the product was extracted at 200 rpm for 1 hour. The water extract was centrifuged at 3,000 rpm for 10 minutes to obtain a supernatant. 2 filter paper was used for α-glucosidase inhibitory activity and DNJ analysis.

Example  2. Results

2.1 DNJ  Biosynthesis Gene group  Isolation and sequencing

In order to isolate the DNJ biosynthesis gene group, primers (MJS-1 and MJS-2) were constructed based on the internal base sequence of the yktc1 gene and PCR was performed using 90 kinds of bacterial genomic DNA isolated from Korean traditional fermented food as a template A DNA fragment of about 470 bp was detected.

For the taxonomic characterization, 16S rRNA gene was analyzed by PCR amplification using 518F and 800R primers. As a result, all selected bacteria were identified as B. amyloliquefaciens (Fig. 1a).

Three sets of primers were constructed to analyze upstream and downstream sequences of the amplified yktc1 gene. The primer set was based on the entire ORF sequence of gabT1 , yktc1 and gutB1 from B. amyloliquefaciens . Sequence analysis of each PCR product revealed that the mutants of GabT1 (422 aa, 4 aminobutylate aminotransferase), Yktc1 (316 aa, inorganic polyphosphate / ATP-NAD kinase) and GutB1 (348 aa, zinc-binding dehydrogenase ) (Table 1).

B. amyloliquefaciens 140N (aa) B. amyloliquefaciens FZB42 (% identity / similarity with aa, 140N) B. atrophaeus 1942 (% identity / similarity with aa, 140N) Expected feature GabTl (422) ABS72608 ^a (425, 98/99) ADP34799 (422, 94/97) 4-aminobutyrate aminotransferase  Yktc1 (316) ABS72609 (316, 96/98) ADP34800 (316, 88/93) Inorganic polyphosphate / ATP-NAD kinase GutB1 (348) ABS72610 (348, 99/100) ADP34801 (348, 86/95) Zinc-binding dehydrogenase

^a B. GenBank accession number of the protein corresponding to each ORF derived from amyloliquefaciens 140N

The homology between the DNJ biosynthesis gene group found in the present invention and the group of genes obtained by combining the genes already known (Fig. 1B and Fig. 2) was examined. The DNJ biosynthesis gene group of B. amyloliquefaciens 140N isolated was B. amyloliquefaciens FZB42 (accession numbers ABS72608-72610), CAU-B946 (CCF03694-03695), IT-45 (EHM06578-06576), and DSM7 (CBI41285-41287) Showed 97-99% identity and 99% similarity with the expected DNJ biosynthesis gene cluster. However, the isolated DNJ biosynthetic gene cluster showed relatively low homology (89% identity and 95% similarity) to B. atrophaeus 1942 (ADP34799-34801). The amino acid sequence of DNJ biosynthesis gene group derived from B. amyloliquefaciens 140N is shown in accession no. I registered with GenBank with JX174443.

2.2  B. amyloliquefaciens  140N DNJ Production of

In order to investigate the relationship between DNJ biosynthesis gene families and DNJ production, the inventors cultured the selected bacteria and measured the alpha-glucosidase inhibitory activity. B. licheniformis 320 without the DNJ biosynthesis gene group was used as a negative control, and the strain showed no inhibitory activity against? -Glucosidase (Table 2).

bacteria α-glucosidase inhibitory activity (%) B. licheniformis 320 0 B. amyloliquefaciens 59N 11.5 B. amyloliquefaciens 432N 24.9 B. amyloliquefaciens 446N 43.9 B. amyloliquefaciens 259N 45.9 B. amyloliquefaciens 140N (KCCM11286P) 78.9

B. amyloliquefaciens 140N showed the highest inhibitory activity (78.9%), and B. amyloliquefaciens 140N was used as a DNJ producing strain, while all the selected bacteria showed α-glucosidase inhibitory activity. UPLC-ESI-Q-TOF-MS analysis (cation mode) of the heat-treated culture supernatant results in a retention time of 5.3 min to confirm the production of DNJ by B. amyloliquefaciens 140N m / z 164.09 A peak corresponding to [M + H] ⁺ was detected (Fig. 3C). The peak was the same as the retention time and the MS spectrum of the DNJ (C ₆ H ₁₃ NO ₄ , calcd 163.08) compound used as a standard product. As a result, the α-glucosidase inhibitory activity of B. amyloliquefaciens 140N was closely related to the biosynthesis of DNJ (Fig. 3G).

2.3 Culture medium and fermentation conditions

B. In order to improve the activity of inhibiting? -Glucosidase by amyloliquefaciens 140N, the carbon source and nitrogen source in the medium were optimized. Except for xylose and galactose, all carbon sources showed an inhibitory activity of? -Glucosidase of 40% or more (FIG. 4). Water soluble starch caused 84.4% inhibition, maltose 80.3% and lactose 62.6%. As a result of using various concentrations of 0 to 3% to find the concentration of water-soluble starch for optimization of? -glucosidase inhibition, the starch concentration gradually increased as the starch concentration increased from 0 to 1%. However, no significant difference was observed at the concentration of 1 ~ 3% and 88.9% of 2% water soluble starch showed the highest inhibition.

To examine the effect of nitrogen source on? -glucosidase inhibitory activity, 1% of various nitrogen sources were added instead of yeast extract (FIG. 5). Malt extract and beef extract showed significantly lower inhibitory activity of α-glucosidase inhibition, while tryptone (96.1%), peptone (93.3%) and yeast extract (91.2%) showed high inhibitory activity. As a result of examining the effect of tryptone concentration on? -Glucosidase inhibitory activity, the tryptone concentration (0.5-3%) did not significantly affect? -Glucosidase inhibitory activity, The activity (96.1%) appeared at 1% tripotone and the inhibitory activity was slightly lowered as the tryptone concentration increased to 3%.

B. amyloliquefaciens 140N was cultured in the presence of 2% water soluble starch, 1% tryptone, 0.05% KH ₂ PO ₄ , and 0.05% (NH ₂ ) ₄ SO ₄ Lt; RTI ID = 0.0 > 37 C < / RTI > using a 5 L fermentor. After 24 hours fermentation, the growth of B. amyloliquefaciens 140N reached the stopper and showed the highest cell growth (1.5 g / L gun cell weight) and DNJ production (0.8 g / L), followed by a similar pattern for 24 h (Fig. 6).

2.4 B. amyloliquefaciens  140N < / RTI > In soybean fermentation DNJ  production

In order to investigate the possibility of developing a fermented food having α-glucosidase inhibitory activity, B. amyloliquefaciens 140N was inoculated to the grown soybeans and fermented at 37 ° C. for 3 days to prepare soybean fermented product. Soybean fermentations prepared with B. licheniformis 320 that did not biosynthesize negative control DNJ showed no inhibition of? -Glucosidase , while soybean fermentations prepared with B. amyloliquefaciens 140N exhibited relatively high? -Glucosidase inhibitory activity (84.4%). As a result of analyzing the soybean fermented product prepared with B. amyloliquefaciens 140N through UPLC-ESI-Q-TOF-MS, the DNJ compound showing the same peak as that of DNJ of the standard product was analyzed. As a result, B. amyloliquefaciens 140N DNJ was also produced during soybean fermentation (Figs. 3E and 3H). However, soybean fermentations prepared by inoculation with B. licheniformis 320 did not have a corresponding peak in UPLC-ESI-Q-TOF-MS (FIG. 3D).

From the above results, B. amyloliquefaciens 140N has a DNJ biosynthesis gene group and can be used to produce functional fermented food for diabetic and obese patients because DNJ having an? -Glucosidase inhibitory activity is biosynthesized. The skilled person will understand. In addition, B. amyloliquefaciens was inoculated with B. amyloliquefaciens 140N (Zhao et al., 2009; Baek et al., 2010), considering that it could be used as a starter of traditional fermented foods together with B. subtilis The crude extract was extracted from the soybean fermented product, and the α-glucosidase inhibitory activity was measured. As a result, the inhibitory activity was as high as 84.4%, and the peak corresponding to DNJ was analyzed in the extract. These results indicate that the B. amyloliquefaciens 140N discovered through the present invention is applicable to the functional food industry having antidiabetic and anti-obesity effects.

Therefore, the present invention can be applied to the production of functional foods or pharmaceuticals capable of DNJ production and α-glucosidase inhibitory activity by B. amyloliquefaciens 140N having the DNJ biosynthesis gene group.

Korea Microorganism Conservation Center (overseas) KCCM11286P 20120614

Claims (17)

Bacillus A method for producing 1- deoxynojirimycin using amyloliquefaciens . The method according to claim 1,
remindBacillus amyloliquefaciensThe Bacillus amyloliquefaciens 140N (KCCM11286P) Method for producing 1-deoxy nojirimycin characterized in that. The method of claim 1,
Deoxynojirimycin is cultured in a culture solution containing at least one carbon source selected from the group consisting of water-soluble starch, maltose, lactose, sucrose and glucose. The method of claim 1,
Wherein the cultivation is carried out in a culture medium containing at least one nitrogen source selected from the group consisting of soytone, casamino acid, peptone, yeast extract, tryptone and whey powder. A method for producing deoxynojirimycin. Bacillus A functional fermented food composition comprising 1- deoxynojirimycin fermented with amyloliquefaciens . 6. The method of claim 5,
Bacillus amyloliquefaciens is Bacillus A functional fermented food composition, characterized in that amyloliquefaciens 140N (KCCM11286P). 6. The method of claim 5,
Functional fermented food composition, characterized in that the fermented food is soybean fermented food. 6. The method of claim 5,
The 1-deoxy nozirimycin is a functional fermented food composition, characterized in that to prevent or alleviate diabetes or obesity through alphaglucosidase inhibitory activity. Bacillus A pharmaceutical composition for preventing or treating diabetes or obesity, comprising 1- deoxynojirimycin fermented with amyloliquefaciens . 10. The method of claim 9,
Bacillus amyloliquefaciens is Bacillus Pharmaceutical compositions, characterized in that amyloliquefaciens 140N (KCCM11287P). 10. The method of claim 9,
The 1-deoxy nozirimycin is a pharmaceutical composition, characterized in that to prevent or treat diabetes or obesity through alphaglucosidase inhibitory activity. Bacillus subject to fermentation A method of producing a functional fermented food comprising 1- deoxynojirimycin , comprising the step of fermenting with amyloliquefaciens . 13. The method of claim 12,
Bacillus amyloliquefaciens is Bacillus amyloliquefaciens 140N (KCCM11286P) Method for producing a functional fermented food, characterized in that. 13. The method of claim 12,
The fermentation target is increased soybean, and the fermented food is a method for producing a functional fermented food, characterized in that the Cheonggukjang. 13. The method of claim 12,
The fermentation is a method for producing a functional fermented food, characterized in that at 30 ℃ to 42 ℃. 13. The method of claim 12,
The 1-deoxy nozirimycin is a method for producing a functional fermented food, characterized in that to prevent or treat diabetes or obesity through alphaglucosidase inhibitory activity. Bacillus 1-deoxynojirimycin producing strain of amyloliquefaciens 140N (KCCM11286P).

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