• Title/Summary/Keyword: Culture broth

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Biochemical Characterization of an Extracellular Xylanase from Aestuariibacter sp. PX-1 Newly Isolated from the Coastal Seawater of Jeju Island in Korea (대한민국 제주도 연안 해수에서 새롭게 분리한 Aestuariibacter sp. PX-1이 생산하는 자일라네이즈의 생화학적 특성)

  • Kim, Jong-Hee
    • Microbiology and Biotechnology Letters
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    • v.48 no.2
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    • pp.215-222
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    • 2020
  • The marine microorganism PX-1, which can hydrolyze xylan, was isolated from coastal sea water of Jeju Island, Korea. Based on the 16S rRNA gene sequence and chemotaxonomy analysis, PX-1 was identified as a species of the genus Aestuariibacter and named Aestuariibacter sp PX-1. From the culture broth of PX-1, an extracellular xylanase was purified to homogeneity through ammonium sulfate precipitation and subsequent adsorption chromatography using insoluble xylan. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration chromatography estimated the molecular weight of the purified putative xylanase (XylA) as approximately 64 kDa. XylA showed xylanase activity toward beechwood xylan, with a maximum enzymatic activity at pH 6.0 and 45℃. Through thin-layer chromatographic analysis of the xylan hydrolysate produced by XylA, it was confirmed that XylA is an endo-type xylanase that decomposes xylan into xylose and xyloligosaccharides of various lengths. The Km and Vmax values of XylA for beechwood xylan were 27.78 mM and 78.13 μM/min, respectively.

Purification and Properties of the Polyvinyl alcohol oxidase from Xanthomonas campestris J2Y (폴리비닐 알콜 분해균 Xanthomonas campestris J2Y의 Polyvinyl alcohol oxidase 정제 및 성질)

  • Kwoen, Dae-Jun;Jo, Youl-Lae
    • Applied Biological Chemistry
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    • v.39 no.5
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    • pp.349-354
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    • 1996
  • The Polyvinyl alcohol(PVA) oxidase involved in PVA degradation by microorganism has been purified to homogeneity from culture broth of Xanthomonas campestris J2Y grown in a minimal medium containing PVA as a sole carbon source. The enzyme was purified by DEAE-cellulose chromatograpy and Sephadex G-150 gel filtration. The purified PVA oxidase was electrophoretically homogeneous both in the absence and presence of SDS. The molecular weight of the enzyme was estimated to be about 55,000 daltons by SDS-polyacrylamide gel electrophoresis and Sephadex G-150 gel filtration. The native enzyme existed as a monomer. The optimal pH and temperature was shown to be pH 7 and $37^{\circ}C$ respectively. The activity of enzyme was stable below $55^{\circ}C$ and between pH range of $5{\sim}11$. The enzyme activity was significantly inhibited by metal compounds such as $Ag^{2+},\;Hg^{2+}$. While, metal ions such as $Mn^{2+},\;and\;Cu^{2+}$ stimulated the reaction. Km value of the enzyme for PVA was $7.04{\times}10^{-2}mmol/{\ell}$.

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EVALUATION OF ENTEROCOCCUS FAECALIS REMOVAL EFFICACY OF THE ENDOVAC® AND ENDOACTIVATOR® INTRACANAL IRRIGATION METHODS (EndoVac®과 EndoActivator®를 이용한 근관세척법의 Enterococcus faecalis 제거 효율 평가)

  • Song, Seung-Gon;Park, Se-Hee;Cho, Kyung-Mo;Kim, Jin-Woo
    • Restorative Dentistry and Endodontics
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    • v.34 no.5
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    • pp.390-396
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    • 2009
  • The aim of this study was to evaluate endodontic irrigation methods with $EndoVac^{(R)}$ and $EndoActivator^{(R)}$ in the elimination of Enterococcus faecalis from the root canals. Extracted 70 human single-rooted teeth were used. The canals were instrumented by a crown-down technique with .04 taper ProFile to ISO size 40. After the teeth were autoclaved, the canals were inoculated with E. faecalis and incubated for 48 h. The teeth were randomly divided into three experimental groups of 20 teeth each according to canal irrigation methods and two control groups as follows: group 1 - $EndoVac^{(R)}$; group 2 - $EndoActivator^{(R)}$; group 3-Conventional needle irrigation method. After canal irrigation using 2.5% NaOCl. first samples (S1) were taken using sterile paper point. And the canals were filled with sterile brain heart infusion (BHI) broth and incubated for 24 h, then second samples (S2) were taken. The samples were cultured on BHI agar plate to determine the numbers of colony forming units (CFU). In first sampling (S1), only one canal of conventional method among the all experimental groups was positive cultured. In second sampling (S2), $EndoVac^{(R)}$ group showed the least positive culture numbers of E. faecalis. There was statistically significant difference between the $EndoVac^{(R)}$ and conventional needle irrigation methods in the mean value of Log CFU. According to the results of this study, $EndoVac^{(R)}$ showed better efficacy than conventional needle irrigation method in the elimination of E. faecalis from the root canal.

Isolation and Phylogenetic Characterization of Chitinase Producing Oligotrophic Bacteria (Chitinase생산 저영양세균의 분리 및 계통분류학적 특성)

  • Kim Soo-Jin;Kim Min-Young;Koo Bon-Sung;Yoon San-Hong;Yeo Yun-Soo;Park In-Cheol;Kim Yoon-Ji;Lee Jong-Wha;Whang Kyung-Sook
    • Korean Journal of Microbiology
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    • v.41 no.4
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    • pp.293-299
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    • 2005
  • Many isolates from soil of Korean ginseng rhizosphere did not show remarkable growth on full strength of the conventional nutrient broth (NB medium) but grew on its 100-fold dilution (DNB medium). Six hundred-forty strains were isolated as oligotrophic bacteria. In the course of screening for new bioactive compounds from oligotrophic bacteria from soil, 8 strains which had appeared to form of clear zone on a medium containing colloidal chitin as a sole carbon source were selected for further studies. Strain CR42 hydrolyzed a fluorogenic analogue of chitin, 4-methylumbelliferyl-D-glucosaminide (MUF-NAG) . Mo st of the culture supernatant of these isolates hydrolyzed 4-methylumbelliferyl-D-N,N'-diacetylchitobioside (MUF-diNAG). The isolates were heterogeneous and categorized to gamma- and beta-proteobacteria, Bacillaceae, Actinobactepia, and Bacteroides by 16S rRNA analysis. Two strains, WR164 and CR18, had a 16S rRNA sequence of $95-96\%$ identical to uncultured bacteria. It was observed that CR2 and CR75 could inhibit the growth of Colletotrichum gloeosporioides with hyphal extention-inhibition assay on PDA plate supplemented with $1\%$ colloidal chitin.

Biological Evaluation of Nargenicin and Its Derivatives as Antimicrobial Anti-inflammatory Agents (토양 균주 발효 추출물 Nargenicin 및 그 유도체의 항생제 대체 효과능 평가)

  • Cho, Seung-Sik;Hong, Joon-Hee;Chae, Jung-Il;Shim, Jung-Hyun;Na, Chong-Sam;Yoo, Jin-Cheol
    • Korean Journal of Organic Agriculture
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    • v.22 no.3
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    • pp.469-481
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    • 2014
  • IIn vitro antimicrobial and anti-inflammatory activities of nargenicin and its derivatives were investigated. Nargenicin, an unusual macrolide antibiotic with potent anti-MRSA (methicilin-resistant Staphylococcus aureus) activity, was purified from the culture broth of Nocardia sp. CS682. And variety of novel nargenicin derivatives was synthesized from nargenicin. Two compounds (4 and 5) exhibit a broad spectrum of antimicrobial activities against infectious bacteria. The antimicrobial activity of derivatives against fifteen organisms was assessed using the minimum inhibitory concentration (MIC). The MIC values were in the ranges of $0.15{\sim}80{\mu}g/mL$ (w/v) for compound 1 and 2, $5{\sim}80{\mu}g/mL$ (w/v) for compound 3, $1.25{\sim}40{\mu}g/mL$ (w/v) for compound 4, and $1.25{\sim}80{\mu}g/mL$ (w/v) for compound 5, depending on the pathogens studied. In vitro, we investigated cytotoxicity and inhibition of nitric oxide (NO) production of synthesized compounds 1-5 in Raw 264.7 cells. LPS-induced nitric oxide releases were significantly blocked by compound 3, 4 and 5 in a dose-dependent manner. At high concentrations ($5{\mu}g/mL$) compound 5 inhibited the NO production by 95%. Compound 4 inhibited the release of NO in LPS-activated Raw 264.7 cells by 75% at the concentration of $10{\mu}g/mL$. Compound 3 inhibited the release of NO in LPS-activated Raw 264.7 cells by 65% at the concentration of $100{\mu}g/mL$. On the other hand, nargenicin, compound 1 and 2 did not inhibit NO production. These results demonstrated that compound 4 and 5 displayed antimicrobial activity and blocked LPS-induced pro-inflammatory mediators such as NO in macrophages, which might be responsible for its therapeutic application.

Bacteriological Study of Listeria sp. Isolated from Seawater and Sea Food (해수와 해산물로부터 Listeria 속의 분리와 세균학적 조사)

  • 강치희;이만효;황용일
    • Journal of Life Science
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    • v.13 no.4
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    • pp.390-399
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    • 2003
  • Four species of the genus of Listeria were isolated from seawater and sea food in Kyungnam province, South Korea. These isolated strains were classified into Listeria sp. from different samples by appropriate cultivation conditions and biochemical tests including serological test. In a day enrichment cultivation, the following strains were found out of 100 samples: L. innocua (35%), L. ivanovii (4%), L. monocytogenes (4%), and L. welshimeri (1%). For seven days enrichment culture, L. innocua (38%), L. ivanoii (5%), L. monocytogenes (7%), and L. welshimeri (1%) were isolated. From these results, Listeria species were more efficiently isolated in seven day enrichment broth than in one day enrichment. However, these isolated Listeria species were less grown in the selective medium than in the enrichment medium. Isolation rates of Listeria species showed differency for each sample and Listeria species were more abundantly isolated in shrimps (80%) and crayfishes (80%) than little neck clams (50%), seawater (25%) and mussels (20%). From the results of serological classes for the seven L. monocytogenes, two strains were defined as type I and the other five strains as type IV.

Microbiological and Immunological Investigation on the Bacteroides gingivalis in Rapidly Progressive and Adult Periodontitis in Korean (한국인 급성진행성 및 성인성 치주염의 원인균인 Bacteroides gingivalis에 대한 미생물 및 면역학적 연구)

  • Chung, Chong-Pyoung;Lee, Jong-Heun;Chung, Hyun-Ju
    • The Journal of the Korean Society for Microbiology
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    • v.22 no.3
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    • pp.309-321
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    • 1987
  • For the investigation of microbiological and immunological specificity of Bacteroides gingivalis, Bacteroides gingivalis were isolated, enumerated and characterized from 13 Korean rapidly progressive periodontitis and 7 healthy control by anaerobic culture technique. The total proportion of black-pigmented Bacteroides from Korean R.P.P. patients and healthy control were 8.78% and 0.92%, respectively, among total isolated black-pigmented Bacteroides. In antibiotic susceptibility test, Bacteroides gingivalis isolated from R.P.P. patients were sensitive to Ampicillin and Tetracycline, and resistant to Gentamicin and Erythromycin in disc diffusion method. In antibiotic broth dilution method, the minimum inhibitory concentration(MIC) to Bacteroides gingivalis was 2 unit/ml of Penicillin and $0.25{\sim}1{\mu}g/ml$ of Tetracycline, respectively. The concentration of serum IgG in rapidly progressive periodontitis patients were sigificantly higher than that of healthy control, and concentration of diluted gingival crevicular IgG has not any significant differences between two groups. Serum and gingival crevicular IgG antibody to Bacteroides gingivalis were significantly higher titer in rapidly progressive periodontitis patients to compare with healthy control. The lipopolysaccharide profiles of 2 Korean B. gingivalis in silver stained sodium dodecyl sulfate-polyacrylamide gel electrophoresis were similar to type strains of B. gingivalis and typical LPS band were appeared around the 24-Kd molecular weight. Immunodiffusion test and immunoelectrophoresis of the L.P.S. extracted from 2 Korean B. gingivalis and 2 kinds of type strains of B. gingivalis showed that B. gingivalis Korean-1 was reacted identically to B. gingivalis ATCC 33277. In trypsin and ${\alpha}$-glucosidase activity test of 2 Korean B. gingivalis, both of them revealed positive trypsin and negative ${\alpha}$-glucosidase activity, respectively. These investigation suggested that B. gingivalis is important pathogenic plaque bacteria for the pathogenesis of periodontitis and further study is needed to purify and characterize of the species-specific antigens of this organisms to develop monoclonal antibody and potential diagnostic reagents.

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Inhibitory Effect of Major Food Components on the Activity of Antimicrobial Active Substance from n-Hexane Fraction of Mallotus japonicus Muell on Listeria monocytogenes (식품의 주요 성분이 예덕나무 헥산추출물의 Listeria monocytogenes에 대한 항균활성에 미치는 영향)

  • Ahn, Yong-Seon;Shin, Dong-Hwa;Kim, Yong-Suk
    • Korean Journal of Food Science and Technology
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    • v.32 no.2
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    • pp.469-476
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    • 2000
  • Inhibitory action of major food components on antimicrobial active substance (linolenic acid) was tested against Listeria monocytogenes ATCC 19111 for 72 hr at $30^{\circ}C$. Linolenic acid (50, 1000 ppm) and n-hexane fraction of Mallotus japonicus Muell (50, 1000 ppm) were added to the broth culture medium containing casein(1%, 3%), soybean oil(1%, 3%) and soluble starch (1%, 3%), respectively. Linolenic acid 1000 ppm and n-hexane fraction 1000 ppm exhibited strongly antimicrobial actions on L. monocytogenes which were not detected viable cell after 24 hr. But casein (3%) and soybean oil (3%) strongly diminished the antimicrobial action of 1000 ppm of n-hexane fraction. Soluble starch (1%, 3%) did not affect the antimicrobial action of 1000 ppm of linolenic acid and n-hexane fraction.

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Effect of Trehalose and Sugar Alcohol on the Viability of Lactic Acid Bacteria and Quality Characteristics during Frozen Storage of Yoghurt (Trehalose와 당류가 냉동요구르트의 저장 중 유산균 생존율과 품질특성에 미치는 영향)

  • Woo, Sung-Ho;Jhoo, Jin-Woo;Yoon, Won-Byong;Kim, Gur-Yoo
    • Food Engineering Progress
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    • v.14 no.1
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    • pp.14-20
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    • 2010
  • The objective of the current study was to determine the cryoprotective effects of trehaolse on lactic acid bacteria in the frozen yoghurt during long-term frozen storage conditions. The frozen yoghurts were prepared using starter culture containing Lactobacillus bulgaricus and Streptococcus thermophilus, as well as trehalose and sorbitol as a cryoprotectant. The viable cell numbers of lactic acid bacteria in frozen yoghurt did not significantly decreased during six weeks frozen storage conditions. The MRS broth, which contains either trehalose or sorbitol, cultured with L. bulgaricus and/or S. thermophilus, and then the cultured medium was kept in the frozen condition for six weeks. The results indicated that lactic acid bacteria viability significantly increased with trehalose addition (2 and 5%) in the media compared to those of control and sorbitol supplement groups. The lactic acid bacteria viability in the yoghurts was examined on the effects of repeated freeze and thaw events. The freeze-thaw resistance of lactic acid bacteria significantly increased with trehalose supplement in the yoghurt. The major volatile aroma compounds (acetaldehyde, acetone, ethanol, diacetyl, and acetoin) in yoghurt were separated and indentified by headspace GC-FID analysis. Distinct flavor components and their ratios are known as important quality factors for yoghurt notes. Trehalose addition to the yoghurt was not influenced these factors during lactic acid fermentation. The results in this study demonstrated that trehalose potentially can be applicable as an effective cryoprotectant for lactic acid bacteria in the frozen yoghurt products.

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.