• Journal of Microbiology and Biotechnology
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• v.22 no.8
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• pp.1092-1100
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• 2012

Ten Bacillus strains with antimicrobial activities were isolated from Cheonggukjang produced at different parts in Korea. They all inhibited Listeria monocytogenes ATCC 19111 and nine inhibited Bacillus cereus ATCC 14579. Four isolates (W42, H27, SKE 12, and K21) showing strong inhibiting activities were identified as B. subtilis. B. subtilis W42 was the most inhibiting strain. The antimicrobial activity of culture supernatant from B. subtilis W42 was destroyed completely by proteinase K treatment, indicating that a bacteriocin was the responsible agent. The bacteriocin, Bac W42, was most stable at pH 7 and stable between pH 3-6 and 8-9. Bac W42 was stable up to $80^{\circ}C$. BHI (brain heart infusion) and TSB (tryptic soy broth) were the best media for the activity (320 AU/ml) followed by LB (160 AU/ml). Bac W42 was partially purified by column chromatographies. The specific activity was increased from 1,151.2 AU/ml to 9,043.5 AU/ml and the final yield was 26.3%. Bac W42 was 5.4 kDa in size as determined by SDS-PAGE. Bac W42 showed bactericidal activity against L. monocytogenes ATCC 19111.

• Applied Biological Chemistry
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• v.50 no.2
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• pp.87-94
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• 2007

A bacterium which has high enzymatic activities such as amylase, cellulase and protease was isolated from Korean traditional soybean food, doenjang. The isolated bacterium was identified to Bacillus subtilis HS25 by the test of morphological and biochemical properties according to Bergey's Manual of Systematic Bacteriology and API 50 CHL kit, and by the 16S rDNA sequence. The isolated B. subtilis HS25 had a potent antibacterial activity against food born causative or pathogenic bacteria. B. subtilis HS25 is endospore forming cell and contained flagella and abundant viscous material at the out layer of cell wall. It was rod type bacterium $(0.5{\sim}0.8{\times}3{\sim}5{\mu}m)$ having biochemical characteristics such as gram staining(+), catalase(+), oxidase(-) and hydrolysis of esculin(+). The optimal medium compositions for production of antibacterial substance in the B. subtilis HS25 were 1% of soluble starch, 0.5% of yeast extract, 0.5% of peptone and 0.05% of MgCl$_2{\cdot}6H_{2}O$. The optimum temperature and pH of the growth of the B. subtilis HS25 was 35$^{\circ}C$ and pH 7.5, respectively. The antibacterial activity was more high in neutral to a little alkaline pH (6.5-10.5) than in acidic pH. The optimal shaking speed to grow and to produce antibacterial substance of the B. subtilis HS25 was 160${\sim}$200 rpm. The optimal culture time for antibacterial activities of the bacterium were shown to be in the range of 12-36 hr.

• The Korean Journal of Food And Nutrition
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• v.26 no.2
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• pp.273-279
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• 2013

The microorganisms producing high protease activity and acid producing ability were isolated from Chunggukjang and kimchi, which were identified as Bacillus subtilis and Lactobacillus planetarum by morphological, biochemical and nutrient requirement. The attempt was made to produce soybean milk yoghurt by using the isolated microorganisms. The mixed culture of Bacillus subtilis and Lactobacillus plantarum exhibited the lowest pH value of 4.23 and highest titratable acidity of 0.88% compared to those of single cultures at $37^{\circ}C$ for 32 hrs, and their total viable count was $4.09{\times}10^8$ $cfu/m{\ell}$. The ${\alpha}$-amylase activity was the highest in culture of Bacillus subtilis after incubation for 24 hrs, while protease activity was most produced in mixed culture of Bacillus subtilis and Lactobacillus plantarum. The amounts of reducing sugars were steadily decreased as soy milk fermentation progressed.

• Journal of Animal Science and Technology
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• v.64 no.2
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• pp.291-301
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• 2022

The objective of this study was to evaluate the effects of different mixing ratios of Bacillus licheniformis and Bacillus subtilis in diets on nutrient digestibility, fecal microflora, and odor gas emissions of growing pigs. A total of four crossbred ([Landrace × Yorkshire] × Duroc) barrows with average body weight (BW) of 41.2 ± 0.7 kg were randomly allotted four diets over four periods in a 4 × 4 Latin square design. Treatments were as follows: Control (CON, basal diet), CON + 0.2% probiotic complex (L4S6, B. licheniformis and B. subtilis at a 4:6 ratio), CON + 0.2% probiotic complex (L5S5, B. licheniformis and B. subtilis at a 5:5 ratio), CON + 0.2% probiotic complex (L6S4, B. licheniformis and B. subtilis at a 6:4 ratio). Dietary probiotic supplementation showed higher crude protein (CP) digestibility values and lower Escherichia coli counts in fecal samples than the CON group (p < 0.05). There was no significant difference in NH₃ or H₂S emission until day 3. The positive effect of H₂S and NH₃ emissions was detected earlier with the L4S6 and L5S5 compared to the L6S4, which had a lower ratio of B. subtilis. Both the L4S6 and L5S5 probiotic complexes significantly decreased the fecal H₂S and NH₃ emission in days 4 and 6 (p < 0.05). On day 7, all probiotic complexes decreased (p < 0.05) H₂S and NH₃ emissions than the CON group. Our results agreed that the dietary supplementation of Bacillus licheniformis and Bacillus subtilis complexes in growing pigs can significantly improve CP digestibility and reduce fecal E. coli counts, NH₃ and H₂S emissions. Notably, the higher mixing ratio of Bacillus subtilis in probiotic supplementation is more effective in reducing the odor of manure.

• Applied Biological Chemistry
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• v.42 no.1
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• pp.20-28
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• 1999

To improve Bacillus strains producing biopolymer, conditions for protoplast formation and regeneration were investigated in biopolymer producing Bacillus subtilis K-1 and lactose utilizing Bacillus coagulans. Bacillus subtilis K-1 mutant (SM-2) and Bacillus coagulans mutants (CM-12) were marked auxotrophic and antibiotics-resistant (SM-2) and an antibiotics-resistant mutants, respectively. To formate protoplasts derived from the mutants, conditions were established as follows. For B. subtilis mutant SM-2, its culture in mid-logarithmic phase was added with penicillin G (1.0 unit/ml) and further reacted for 1.5 hr. Cells were collected and then treated in lysis fluid (pH 7.0) containing 0.4 M sucrose and lysozyme $25\;{\mu}g/ml$ for 40 min at $37^{\circ}$. Protoplast formation was very successful (99.6%) and the ratio of cell wall regeneration was 2.4%. For Bacillus coagulans mutant CM-12, its mid-logarithmic phase culture was treated with penicillin G (0.3 unit/ml) and glycine (0.5%) for 1hr. Cells were collected and then resuspended in lysis buffer (pH 7.0) containing 0.6 M lactose and lysozyme $(300\;{\mu}g/ml)$ for 30 min at $37^{\circ}$. Protoplast formation was also successful (90.8%) and cell wall regeneration ratio was similar to SM-2 (2.2%). To improve regeneration frequency, regeneration medium was obtained as followed condition,. Cell wall regeneration was improved 2-4 folds with 5.1% for B. subtilis SM-2 and 10.3% for B. coagulans CM-12 when protoplasts mixed with soft top agar(0.4%) was overlaid onto trypticase soy broth medium containing 0.4 M sucrose, 0.7% casamino acid, 1% PVP, 25 mM $MgCl_2,\;25\;mM\;CaCl_₂$ and 1.5% agar.

• Korean Journal of Microbiology
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• v.43 no.4
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• pp.331-336
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• 2007

A bacterium producing the alkaline pretense was isolated from Chungkookjoug, and was identified as Bacillus subtilis JK-1 based on morphological, physiological and biochemical characteristics, as well as phylogenetic analysis using 16S rRNA gene sequence. The optimum pH and temperature of the pretense activity were pH 9.0 and $55^{\circ}C$, respectively. This enzyme was stable at the temperatures $40{\sim}80^{\circ}C$. The maximum alkaline pretense production was obtained when 1.0% (w/v) xylose, 1.0% (w/v) yeast extract and 0.3% (w/v) $CuSO_4$ were used as carbon source, nitrogen source and mineral source. Under the optimal condition, growth of the isolate was reached at stationary phase after 12 hr followed by incubation, the alkaline pretense production reached a maximum level with $16{\sim}20$ hr cultivation.

• Food Science and Preservation
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• v.15 no.2
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• pp.300-305
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• 2008

We investigated an antibacterial substance produced by Bacillus subtilis HS 25. Antibacterial activity was relatively heat-stable, with no effect caused by heating at $100^{\circ}C$ for 10 min, but a gradual decrease in activity after 15 min at $100^{\circ}C$. The antibacterial substance was more stable at pH 7.42-12 than at pH 4.5-5.0. There was strong antibacterial activity against E. coli and P. mirabilis at pH 12. The minimum inhibition concentration (MIC) of the substance was 5 mg/mL for E. coli 7.5 mg/mL for P. mirabilis and S. aureus and 15 mg/mL for S. enteritidis, K. pneunoniae, and V. parahaemolyticus. When the substance was added to cultures of E. coli, S. enteritidis, and P. mirabilis, the bacterial surfaces became irregular and deeply changed. The substance produced from B. subtilis HS 25 was not degraded by Papain but was degraded by a protease from Aspergillus orzae, pancreatin, and pepsin.

• Research in Plant Disease
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• v.12 no.2
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• pp.85-90
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• 2006

Bacillus subtilis BD0310 isolated from tea leaves was used for the development of a biofungicide against Pestalotiopsis longiseta causing gray blight of tea plants. The optimum growth conditions were investigated for the mass cultivation of the microbial agent. The optimum temperature and cultivation time were determined as $12{\sim}24$ hours at $30^{\circ}C$ and the optimum initial pH was pH 7.0 in nutrient broth. Among the tested carbon sources of fructose, galactose, glucose, glycerol, inositol, lactose, maltose, sorbitol and starch, maltose and inositol were found to highly increase antifungal activity of the microbial agent against P. longiseta. Yeast extract and tryptone apparently increased antifungal activity of the microbial agent among the tested nitrogen sources of casein, tryptone, malt extract, yeast extract and $(NH_4)_2SO_4$. The results will make a contribution to mass production of the antagonistic bacterium Bacillus subtilis BD0310 for development of a microbial agent controlling gray blight of tea plants.

• Microbiology and Biotechnology Letters
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• v.26 no.6
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• pp.507-514
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• 1998

The strain K-54, the best producer of fibrinolytic enzyme, was isolated from Korean traditional food Chung Guk Jang and identified as Bacillus subtilis. Fibrinolytic enzyme was purified and characterized, and its molecular weight was determined. The fibrinolytic enzyme activity was increased about 66.9 times via purification with recovery yield of 10.1%. The optimum pH and temperature of this enzyme were 11 and $65^{\circ}C$. The enzyme was stable within a pH range 8-12 and unstable at 9$0^{\circ}C$. The molecular weight was estimated to be 29,000 dalton in the form of monomer with no other subunit. The isoelectric point was calculated 8.67. N-terminal sequence was identified Ala-Gly-Ser-Val-Pro-Try-Gly-Ser. Km value of the enzyme for $\alpha$-casein was calculated to be 0.31 (3.1 mg/$m\ell$). The enzyme activity highly inhibited by PMSF at 1 nM.

• Applied Biological Chemistry
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• v.46 no.2
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• pp.69-73
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• 2003

A novel antifungal antibiotic for azole-resistant Candida albicans was purified from the culture broth of Bacillus subtilis LAM 97-44 by butanol extraction, Diaion HP-20 and Dowex-50 adsorption chromatography, silica gel flash chromatography followed by HPLC and designated LAM-44A. LAM-44A was stable for 60 min at $100^{\circ}C$, and pH range from 2 to 10. MIC values were observed at $0.5-3.5\;{\mu}g/ml$ against various Candida albicans strains. The antibiotic showed no cytotoxicity for S180, MKN-45, P388, HeLa and 373 at the concentration of 1 mg/ml. LAM-f4A was colorless powder soluble in water, methanol, ethanol, butanol and negative to ninhydrin reaction. The antibiotic had maximum absorption at 273 nm in methanol, and melting point was $202^{\circ}C$. The molecular weight and formula were determined to be 282 and $C_{14}H_{34}O_5$ by $^1H-NMR,\;^{13}C-NMR$, IR spectrum and elemental analysis.