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Purification and Biochemical Characterization of β-agarase Produced by Marine Microorganism Cellulophga sp. J9-3 (해양미생물 Cellulophga sp. J9-3이 생산하는 베타-아가레이즈의 분리 및 생화학적 특성)

  • Kim, Da Som;Kim, Jong-Hee;Chi, Won-Jae
    • Microbiology and Biotechnology Letters
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    • v.49 no.3
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    • pp.329-336
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    • 2021
  • Cellulophga sp. J9-3, is a gram-negative, aerobic marine bacterium belonging to the family Flavobacteriaceae. In addition to cellulose degradability, the J9-3 strain is also capable of hydrolyzing agar in the solid and liquid medium, and the production of agarase in the presence of agarose can be remarkably induced by the bacterium. From the cell culture broth of Cellulophga sp. J9-3, ammonium sulfate precipitation and three kinds of column chromatography were successively performed to purify a specific agarase protein, the AgaJ93. Purified AgaJ93 showed the strongest hydrolyzing activity towards agarose (approximately 22%), and even displayed activity towards starch. AgaJ93 hydrolyzed agarose into neoagarotetraose and neoagarohexaose via various oligosaccharide intermediates, indicating that AgaJ93 is an endo-type β-agarase. AgaJ93 showed maximum activity at a pH of 7.0 and temperature of 35 ℃. Its activity increased by more than six times in the presence of Co2+ ions. The N-terminal sequence of AgaJ93 showed 82% homology with the heat-resistant endo-type β-agarase Aga2 of Cellulophaga sp. W5C. However, the biochemical properties of the two enzymes were different. Therefore, AgaJ93 is expected to be a novel agarose, different from the previously reported β-agarases.

Effect of pH on Growth and Ethanol Production of Clostridium ljungdahlii (Clostridium ljungdahlii 성장 및 에탄올 생산에 pH가 미치는 영향)

  • Park, So Jung;Hong, Sung-Gu;Kang, Kwon-Kyoo;Kim, Young-Kee
    • Applied Chemistry for Engineering
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    • v.22 no.5
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    • pp.562-565
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    • 2011
  • In this study, we developed a bioprocess using Clostridium ljungdahlii as a biological catalyst to produce bio-ethanol, and the effect of pH on microbial growth and ethanol production was investigated. From the results of fermentation at various initial pH condition without pH control, pH of fermentation broth decreased to 4.5 within 24 h due to accumulation of by-product acetic acid and both microbial growth and ethanol production were stopped. The experimental result of initial pH 8 showed the highest microbial growth and ethanol production (0.53 g/L), since the pH drop was relatively slow. From the experiment of pH 7 maintained fermentation using pH controllable bioreactor, the maximum cell dry weight of 1.65 g/L and the maximum ethanol concentration of 1.43 g/L were obtained within 24 h. In conclusion, the C. ljungdahlii growth was enhanced by pH maintenance of neutral range, and the ethanol production was also enhanced based on the growth-associated ethanol production characteristics of C. ljungdahlii.

Red fluorescence of oral bacteria is affected by blood in the growth medium (성장배지 혈액 유무가 구강미생물의 적색 형광 발현에 미치는 영향)

  • Jeong, Seung-Hwa;Yang, Yong-Hoon;Lee, Min-Ah;Kim, Se-Yeon;Kim, Ji-Soo
    • Journal of Korean Academy of Oral Health
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    • v.41 no.4
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    • pp.290-295
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    • 2017
  • Objectives: Dental plaque emits red fluorescence under a visible blue light near the ultra-violet end of the light spectrum. The fluorescence characteristics of each microorganism have been reported in several studies. The aim of this study was to evaluate changes in red fluorescence of oral microorganisms that is affected by blood in the culture media. Methods: The gram-positive Actinomyces naeslundii (AN, KCTC 5525) and Lactobacillus casei (LC, KCTC 3109) and gram negative Prevotella intermedia (PI, KCTC 3692) that are known to emit red fluorescence were used in this study. Each bacterium was activated in broth and cultivated in different agar media at $37^{\circ}C$ for 7 days. Tryptic soy agar with hemin and vitamin $K_3$ (TSA), TSA with sheep blood (TSAB), basal medium mucin (BMM) medium, and BMM with sheep blood (BMMB) were used in this study. Fluorescence due to bacterial growth was observed under 405-nm wavelength blue light using the quantitative light-induced fluorescence-digital (QLF-D) device. The red, green, and blue fluorescence values of colonies were obtained using image-analysis software and the red to green ratio (R/G value) and red to total RGB ratio (R/RGB value) were calculated for quantitative comparison. Results: The QLF-D images of the AN, LC, and PI colonies showed red fluorescence in all media, but the fluorescence of all bacteria was reduced in TSA and BMM media, compared with in TSAB and BMMB media. Both the R/G and the R/RGB values of all bacteria were significantly reduced in growth media without blood (P<0.001). Conclusions: Based on this in vitro study, it can be concluded that red fluorescence of oral bacteria can be affected by growth components, especially blood. Blood-containing medium could be a significant factor influencing red fluorescence of oral bacteria. It can be further hypothesized that bleeding in the oral cavity can increase the red fluorescence of dental plaque.

Isolation of a Pseudoalteromonas sp. JH-1 Producing Agarase and Characterization of its Agarase (Agarase를 생산하는 Pseudoalteromonas sp. JH-1의 분리·동정 및 agarase의 특성 연구)

  • Lee, Dong-Geun;Kim, Ju-Hui;Lee, Sang-Hyeon
    • Journal of Life Science
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    • v.31 no.5
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    • pp.496-501
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    • 2021
  • In this study, the marine agar-degrading bacterium Pseudoalteromonas sp. JH-1 was isolated, and its growth and agarase properties were investigated. Seawater was collected from the offshore of the Yonggung Temple in Busan, and agar-degrading bacteria were isolated and cultured with marine agar medium. The bacterium Pseudoalteromonas sp. JH-1 was isolated through 16S rRNA gene sequencing. The extracellularly secreted enzyme was obtained from the culture broth of Pseudoalteromonas sp. JH-1 and was used to characterize its agarase. The extracellular agarase exhibited a maximum activity of 116.6 U/l at 50℃ and pH 6.0 of 20 mM Tris-HCl buffer. Relative activities were 31, 59, 94, 100, 45, and 31% at 20, 30, 40, 50, 60, and 70℃, respectively. Relative activities were 49, 85, 100, 86, 81, and 67% at pH 4, 5, 6, 7, 8, and 9, respectively. Residual activity was more than 85% after exposure at 20, 30, and 40℃ for 2 hr, and more than 82% after exposure at 50℃ for 2 hr. Zymogram analysis confirmed that Pseudoalteromonas sp. JH-1 produced at least two agarases of 55 and 97 kDa. As the products of α-agarase and β-agarase have antioxidation, antitumor, skin-whitening, macrophage activation, and prebiotic effects, further studies are needed on the agarase of Pseudoalteromonas sp. JH-1.

Optimal Conditions for the Production of Gamma-aminobutyric Acid by Enterococcus casseliflavus PL05 Isolated from Oenanthe javanica

  • Choi, Se Mi;Kim, Jeong A;Kim, Geun Su;Kwon, Do Young;Kim, Sang Gu;Lee, Sang yun;Lee, Kang Wook
    • Journal of Food Hygiene and Safety
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    • v.37 no.1
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    • pp.21-28
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    • 2022
  • In this study, a new lactic acid bacterium (LAB) that could produce gamma-aminobutyric acid (GABA) was isolated from Oenanthe javanica (water celery) and identified as an Enteroccoccus casseliflavus strain. Until recently, there have been many studies on the gamma-aminobutyric acid producing lactic acid bacterium, as well as on some lactic acid bacterium in Enteroococcs genus, but none on the species E. casseliflavus. Therefore, in the purpose of finding the optimal conditions for GABA production of E. casseliflavus PL05, the effects of several conditions including the type of mediums, growth temperatures, initial pH, growth time, L-mono sodium glutamate (MSG) concentration, and carbon source were tested. The study revealed that the PL05 strain grew better in the Brain Heart Infusion (BHI) medium than in the Man, Rogosa, and Sharpe (MRS) or Tryptic Soy Broth (TSB) medium. Also, similar results were obtained with GABA production conditions. As a result of analysis on the GABA production yield by concentration of MSG, a GABA substrate, the highest production was found at 7% of MSG concentration. However, since similar level of production was found at 5%, it is considered to be more efficient to use 5% MSG concentration. The analysis on the growth and GABA production yield by carbon sources showed the highest results when maltose was used. From the final test under the optimal conditions found, 140.06±0.71 mM of GABA was produced over 24 hours with the conversion rate of 78.95%. Lastly, from the sensitivity analysis on the 10 different antibiotics, including vancomycin, it was found that there were not confirmed cases of resistance.

The antimicrobial activity of Daehwanggo against Methicillin-resistant Staphylococcus aureus (대황고(大黃膏)의 메티실린 내성 황색 포도상구균에 대한 항균활성)

  • Lee, Sun-Ae;Kong, Ryong;Kang, Ok-Hua;Seo, Yun-Soo;Zhou, Tian;Kim, Sang-A;Song, Ok-Hee;Kim, Min-Chul;Han, Hyoung-Sun;Choi, Ji-Na;Lee, Young-Seob;Kwon, Dong-Yeul
    • The Korea Journal of Herbology
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    • v.32 no.2
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    • pp.87-95
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    • 2017
  • Objectives : Infectious diseases by Methicillin-Resistant Staphylococcus aureus (MRSA) are a growing problem worldwide. Characteristic of MRSA is endlessly mutation to resist antibiotics. Daehwanggo (DHG) is one of the oriental medicine prescriptions contained in Principles and Practice of Eastern Medicine. Daehwanggo was mainly used for external preparation from old times. The purpose of this study is to confirm possibility as supplementary drug of DHG about antibiotics through observation of synergy effect between DHG and commercial antibiotics and to observe restriction on growth of MRSA on any pathway through observation of mechanism. Methods : The minimum inhibitory concentration (MIC) of DHG against MRSA is $500{\sim}2000{\mu}g/m{\ell}$ by broth dilution method. In the checkerboard method, the combinations of DHG with antibiotics has partial synergistic effect or synergy effect and DHG markedly reduced the MICs of the antibiotics oxacillin (OX), gentamicin (GT) against MRSA. In the inhibition of resistance mechanism of DHG against MRSA, the expression of resistance gene and protein about ${\beta}-lactam$ antibiotic was reduced. Also, we observed the effect of DHG about cell membrane permeability against MRSA, and confirmed that DHG suppressed growth of strains by increasing cell membrane permeability. Results : Basis on the result, we speculate that DHG increase antibacterial activity of antibiotics against MRSA by changing the structure of cell wall of MRSA. Conclusions : These data suggest that Daehwanggo possesses possibility as supplementary drug about antibiotics against MRSA.

Characterization of exopolysaccharide-producing lactic acid bacteria from Taiwanese ropy fermented milk and their application in low-fat fermented milk

  • Ng, Ker-Sin;Chang, Yu-Chun;Chen, Yen-Po;Lo, Ya-Hsuan;Wang, Sheng-Yao;Chen, Ming-Ju
    • Animal Bioscience
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    • v.35 no.2
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    • pp.281-289
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    • 2022
  • Objective: The aim of this study was to characterize the exopolysaccharides (EPS)-producing lactic acid bacteria from Taiwanese ropy fermented milk (TRFM) for developing a clean label low-fat fermented milk. Methods: Potential isolates from TRFM were selected based on the Gram staining test and observation of turbid suspension in the culture broth. Random amplified polymorphic DNA-polymerase chain reaction, 16S rRNA gene sequencing, and API CHL 50 test were used for strain identification. After evaluation of EPS concentration, target strains were introduced to low-fat milk fermentation for 24 h. Fermentation characters were checked: pH value, acidity, viable count, syneresis, and viscosity. Sensory evaluation of fermented products was carried out by 30 volunteers, while the storage test was performed for 21 days at 4℃. Results: Two EPS-producing strains (APL15 and APL16) were isolated from TRFM and identified as Lactococcus (Lc.) lactis subsp. cremoris. Their EPS concentrations in glucose and lactose media were higher than other published strains of Lc. lactis subsp. cremoris. Low-fat fermented milk separately prepared with APL15 and APL16 reached pH 4.3 and acidity 0.8% with a viable count of 9 log colony-forming units/mL. The physical properties of both products were superior to the control yogurt, showing significant improvements in syneresis and viscosity (p<0.05). Our low-fat products had appropriate sensory scores in appearance and texture according to sensory evaluation. Although decreasing viable cells of strains during the 21-day storage test, low-fat fermented milk made by APL15 exhibited stable physicochemical properties, including pH value, acidity, syneresis and sufficient viable cells throughout the storage period. Conclusion: This study demonstrated that Lc. lactis subsp. cremoris APL15 isolated from TRFM had good fermentation abilities to produce low-fat fermented milk. These data indicate that EPS-producing lactic acid bacteria have great potential to act as natural food stabilizers for low-fat fermented milk.

The Growth-Promoting Effect of Pomegranate Concentrates on Lactic Acid Bacteria and Their Application to Yogurt (석류 농축액의 유산균에 대한 성장촉진효과와 요구르트의 적용)

  • Yun Jeong Go;Woan Sub Kim
    • Journal of Dairy Science and Biotechnology
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    • v.41 no.2
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    • pp.76-85
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    • 2023
  • This study investigated the effect of the addition of pomegranate concentrate to yogurt on the growth of pathogenic and lactic acid bacteria. The concentration of the MRS broth was adjusted to one-half and used for an experiment. Pomegranate concentrate was added at concentrations of 4%, 2%, 1%, and 0.5%, which significantly promoted the growth of Lacto-coccus cremoris, Weissella cibaria, Weissella paramesenteroides, Lactobacillus plantarum, Lactobacillus acidophilus, Streptococcus thermophillus, Lactobacillus bulgaricus, and Lactobacillus lactis. The growth of lactic acid bacteria increased with higher concentrations of pomegranate. However, the addition of pomegranate concentrate inhibited the growth of Escherichia coli KCCM11587, E. coli KCCM11591, E. coli KCCM11596, and E. coliKCCM11600. Yogurt with added pomegranate concentrate demonstrated optimal conditions compared to that of the control without the addition. Particularly, the viable cell count of lactic acid bacteria was significantly higher in the yogurt with pomegranate concentrate. Furthermore, the viability of the lactic acid bacteria in the yogurt with pomegranate concentrate was higher than that of the control without the addition of concentrate during storage.

The A Literary Investigation on Mandu (Dumpling);Types and Cooking Methods of Mandu (Dumpling) During the Joseon Era (1400's${\sim}$1900's) (만두의 조리방법에 대한 문헌적 고찰;조선시대 만두의 종류와 조리방법에 대한 문헌적 고찰(1400년대${\sim}$1900년대까지))

  • Bok, Hye-Ja
    • Journal of the Korean Society of Food Culture
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    • v.23 no.2
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    • pp.273-292
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    • 2008
  • Among all the ingredients usedin mandu, the following types were used:, 13 types of grains were used (12.38%), 30 types of vegetables, fruits, bulbs,and nuts were used (28.57%), 32 types of marine products, birds, meats, fishes, and shellfishes were used (30.48%), 10 types of functional ingredients were used (9.52%) and. For spices, 20 types of spices were used (19.05%). 2. Cooking Methods offor Mandu. The mMandu eaten at in the early Joseon era had was primarily made ofusedbuckwheat that contained boiled tofu or egg uiijuk in the kneaded dough for the most part and while kneading with buckwheat, the tofu or egg uiijuk has been boiled down to knead the dough, and and starch powder, bean powder, or rice powder, etc were mixed to make the mandu coating. Buckwheat powder was mixed toadded to the flourwer or was used by itself, while meat, vegetables, tofu, and shiitake mushroom, etc were also addedincluded. From the 18th century, the host plant, or cabbage kimchi, were prepared and combined had been sliced to be used as filling together while red pepper powder was mixed combined withto spices or vinegar soy sauce to be used together. Also, Radishes had beenwere also used as filling, but shown as not being used fromafter the start of the 1900's. For the shape of mMandu, it was madeinto different shapes such as as triangle, rectangle, date plum, gwebul, half moon, or pomegranate shapes, and then shapes to be boiled in simmering water, baked, or cooked as soup in clear broth for soup., In the 17th to 18th century, boilingthen in a steamer gradually became a cooking style, assumed the style of boiling in a steamer in $17th{\sim}18th$ century while in the 16th century,the an essay ofn fermenting flour in ‘Food Dimibang’ in 16th century had indicated it was cooked as the style ofby steaming in a rice steamer. Also, Mandu may have also contained the following: the thin-cut and boiled fish was cut out thin to put into the filling and boiled down, made by putting in added pine nuts after making bbeef jerky or boiled- down meat, fish, or shellfish itself to extractsand mold mandu only the ingredients combined withto put on starch powder, and then boiled down and put on pine nut powder finally, after it or cooled it wasdown to be eaten by dipping in vinegar soy sauce. In conclusion, many different types of mandu were made during the Joseon era using a variety ofwhile the ones using such various ingredients. are also one type of mandu.

Distribution of Beneficial Bacteria in the Intestines after Enzamin Ingestion of Bacillus subtilis AK Strain Fermentation (Bacillus subtilis AK균 발효액(Enzamin)의 섭취 후 장내 유익세균의 분포조사)

  • Ryu, Seo Won;Lee, Hyung H.
    • Journal of Naturopathy
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    • v.7 no.2
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    • pp.27-38
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    • 2018
  • Purpose: The purpose of this study was to investigate whether intestinal proliferation is promoted in beneficial intestinal bacteria or decreased in harmful bacteria before and after ingesting Bacillus fermentation broth (ENM) for 8 weeks in the 16 subjects. Method: Intestinal bacteria were identified by PCR amplification using specific 16S rRNA primers. Results: The Bifidobacterium gene index(%)(gi%) increased to 58.92% in the control group and 69.53% in the test group after the ingestion of ENM, but there was no significant difference. Lactobacillus gi% increased significantly (49.37% in the control and 66.43% in the test) (p<.029). Clostridium gi% was significantly decreased after treatment (83.16% in the control and 67.76% in the test) (p<.077). Bacteroides gi% increased significantly (12.58% in the control and 20.87% in the test) after ingesting (p<.095). Prevotella gi% increased significantly (7.55% in the control and 17.28% in the test) after ingesting (p<.005). After ingesting, the median bacteria increased significantly in the control (20.06%) and the test (35.88%) (p<.001). Conclusions: After ingestion of the ENM, the number of beneficial bacteria increased and the number of harmful bacteria Clostridium tended to decrease. This suggests that ingestion of the Bacillus fermented beverage ENM has an effect on the proliferation of intestinal bacteria.

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