• Title/Summary/Keyword: bacterial enzymes

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A Study on Anti-inflammatory and Antioxidant Effect of Fraxinus rhynchophylla Using Carbohydrate-Hydrolyzing Enzymes (당분해효소를 이용한 물푸레나무 효소처리물의 항염 및 항산화 효능 연구)

  • Hye Won Lee;You Ah Kim;Byoung Jun Park;Sang Keun Han
    • Journal of the Society of Cosmetic Scientists of Korea
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    • v.49 no.3
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    • pp.277-283
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    • 2023
  • Fraxinus rhynchophylla Hance (F. rhynchophylla) is a traditional medicinal plant that has been widely used in East Asia and has been used for chronic bronchitis, bacterial dysentery and improved eyesight. F. rhynchophylla contains various type of coumarins such as esculin, esculetin, fraxin and fraxetin. Esculetin possesses versatile activities including antioxidant, anti-inflammatory, antimicrobial, anticancer properties and improvement of atopic dermatitis. However, there is no research on the process of increasing active components in F. rhynchophylla. The objectives of the present study were to apply biotransformation technology to F. rhynchophylla for increasing the content of esculetin, and enhancing anti-inflammatory and antioxidant activities. F. rhynchophylla extract (FRE) treated with viscozyme L (FRE-VL) showed 3.1 times higher content of esculetin than FRE, and exhibited effects such as increased anti-inflammatory activity and DPPH radical scavenging activity. Based on the these results, it is concluded that biotransformed FRE-VL could be potentially applicable as a new active ingredient in the cosmetic field.

Isolation and biochemical characterization of acid tolerance xylanase producing Bacteria, Bacillus sp. GJY from city park soil (도심공원으로부터 산내성 xylanase를 생산하는 박테리아 분리 및 효소학적 특성)

  • Jang, Min-Young;Park, Hwa Rang;Lee, Chong Gyu;Choo, Gab-Chul;Cho, Hyun Seo;Park, Sam-Bong;Oh, Ki-Cheol;Kim, Bong-Gyu
    • Journal of Applied Biological Chemistry
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    • v.60 no.1
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    • pp.79-86
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    • 2017
  • Microbes in forest are very important due to not only to enhance soil fertility but also maintain a healthy ecosystem by supplying the energy available to living organisms by producing various kinds of enzymes related to degradation of lignocellulosic biomass. In order to isolate a lignocellulosic biomass degrading bacterial strain from the Jurassic park located in Gyeongnam National University of Science and Technology, We used the Luria-Bertani-Carboxymethyl cellulose (CMC) agar trypan blue method containing 0.4 % carboxymethyl cellulose and 0.01 % trypan blue. As a result, we isolated a bacterial strain showing both activity on the CMC and xylan. To identify the isolated strain, 16S rRNA sequencing and API kit analysis were used. The isolated strain turned out to belong to Bacillus species and then named Bacillus sp. GJY. In the CMC zymogram analysis, it showed that one active band of about 28kDa in size is present. Xylan zymogram analysis also showed to have one active band of about 25kDa in size. The optimal growth temperature of Bacillus sp. GJY was $37^{\circ}C$. The maximal activities of CMCase and xylanase were 12 hour after incubation. The optimal pH and temperature for CMCase were 5.0 and $40^{\circ}C$, respectively, whereas the optimal pH and temperature for xylanase was 4.0 and $40^{\circ}C$. Both activities for CMCase and xylanase showed to be thermally stable at 40and $50^{\circ}C$, while both activities rapidly decreased at over $60^{\circ}C$.

Anti-inflammatory Activity of Antimicrobial Peptide Zophobacin 1 Derived from the Zophobas atratus (아메리카왕거저리 유래 항균 펩타이드 조포바신 1의 항염증활성)

  • Shin, Yong Pyo;Lee, Joon Ha;Kim, In-Woo;Seo, Minchul;Kim, Mi-Ae;Lee, Hwa Jeong;Baek, Minhee;Kim, Seong Hyun;Hwang, Jae Sam
    • Journal of Life Science
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    • v.30 no.9
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    • pp.804-812
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    • 2020
  • The giant mealworm beetle, Zophobas atratus (Coleoptera: Tenebrionidae) has been used as a protein source for small pets and mammals. Recently, it was temporarily registered in the list of the Food Code. We previously performed an in silico analysis of the Zophobas atratus transcriptome to identify putative antimicrobial peptides and identified several antimicrobial peptide candidates. Among them, we assessed the antimicrobial and anti-inflammatory activities of zophobacin 1 that was selected bio-informatically based on its physicochemical properties against microorganisms and mouse macrophage Raw264.7 cells. Zophobacin 1 showed antimicrobial activities against microorganisms without inducing hemolysis and decreased the nitric oxide production of the lipopolysaccharide-induced Raw264.7 cells. Moreover, ELISA and Western blot analysis revealed that zophobacin 1 reduced expression levels of pro-inflammatory enzymes such as inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). We also investigated expression of pro-inflammatory cytokines (interleukin-6 and interleukin-1β) production through quantitative real time-PCR and ELISA. Zophobacin 1 markedly reduced the expression level of cytokines through the regulation of mitogen-activated protein kinases (MAPKs) and nuclear factor kappa B (NF-κB) signaling. We confirmed that zophobacin 1 bound to bacterial cell membranes via a specific interaction with lipopolysaccharides. These data suggest that zophobacin 1 could be promising molecules for development as antimicrobial and anti-inflammatory therapeutic agents.

Isolation of Agarivorans sp. KC-1 and Characterization of Its Thermotolerant β-Agarase (한천분해세균 Agarivorans sp. KC-1의 분리 및 내열성 β-아가라제의 특성 규명)

  • Min, Kyung-Cheol;Lee, Chang-Eun;Lee, Dong-Geun;Lee, Sang-Hyeon
    • Journal of Life Science
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    • v.28 no.9
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    • pp.1056-1061
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    • 2018
  • This article reports an agar-degrading marine bacterium and characterizes its agarase. The agar-degrading marine bacterium, KC-1, was isolated from seawater on the shores of Sacheon, in Gyeongnam province, Korea, using Marine Broth 2216 agar medium. To identify the agar-degrading bacterium as Agarivorans sp. KC-1, phylogenetic analysis based on the 16S rRNA gene sequence was used. An extracellular agarase was prepared from a culture medium of Agarivorans sp. KC-1, and used for the characterization of enzyme. The relative activities at 20, 30, 40, 50, 60, and $70^{\circ}C$ were 65, 91, 96, 100, 77, and 35%, respectively. The relative activities at pH 5, 6, 7, and 8 were 93, 100, 87, and 82%, respectively. The extracellular agarase showed maximum activity (254 units/l) at pH 6.0 and $50^{\circ}C$ in 20 mM of Tris-HCl buffer. The agarase activity was maintained at 90% or more until 2 hr exposure at $20^{\circ}C$, $30^{\circ}C$ and $40^{\circ}C$, but it was found that the activity decreased sharply from $60^{\circ}C$. A zymogram analysis showed that Agarivorans sp. KC-1 produced 3 agar-degrading enzymes that had molecular weights of 130, 80, and 69 kDa. A thin layer chromatography analysis suggested that Agarivorans sp. KC-1 produced extracellular ${\beta}$-agarases as it hydrolyzed agarose to produce neoagarooligosaccharides, including neoagarohexaose (21.6%), neoagarotetraose (32.2%), and neoagarobiose (46.2%). These results suggest that Agarivorans sp. KC-1 and its thermotolerant ${\beta}$-agarase would be useful for the production of neoagarooligosaccharides that inhibit bacterial growth and delay starch degradation.

Expression of Matrix Metalloproteinase-9 and Tissue Inhibitor of Metalloproteinase-1 after Administration of Endotoxin in Diabetic Rats (내독소로 자극된 당뇨 쥐에서 단백분해효소와 그 억제제 발현)

  • Seo, Ki Hyun;Choi, Jae Sung;Na, Joo Ok;Uh, Soo Taek;Kim, Yong Hoon;Park, Choon Sik
    • Tuberculosis and Respiratory Diseases
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    • v.61 no.3
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    • pp.256-264
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    • 2006
  • Background: An acute lung injury(ALI) is characterized by the recruitment, activation, and apoptosis of inflammatory cells, numerous products released by inflammatory cells such as reactive oxygen species, inflammatory mediators, and a variety of proteolytic enzymes. It was reported that bacterial infections in diabetics showed impaired PMN functions such as reduced PMN respiratory burst and decreased microbicidal activity in inflamed tissue. However, the effect of the proteinase - inhibitor (MMP-9 vs TIMP-1) in ALI in diabetics is unclear. This study evaluated the differences in the expression of MMP-9 and TIMP-1 after the stimulation of endotoxin in a rat model. Methods: Six-week-old male Sprague-Dawley rats were classified into normal, DM, LPS and DM+LPS groups. The peripheral blood, BAL fluids, and lung tissues were obtained from individual rats. The MMP-9 activity was measured by gelatin zymography and the TIMP-1 level was measured by Western blotting. Results: The total BAL cells of the DM-LPS groups were significantly lower than the LPS groups (p < 0.01). The MMP-9 activities in the serum were higher in the DM+LPS groups than in the other groups. The MMP-9 activities in the BAL fluids were significantly higher in the DM+LPS group than in the normal and diabetic rats (p < 0.05). TIMP-1 expressions in the BAL fluids were significantly lower in the DM+LPS group than other groups (p < 0.05). The ratio between MMP-9 and TIMP-1 in the BAL fluids was significantly higher in the DM+LPS groups (p < 0.05). Conclusion: In ALI in diabetics the higher MMP-9 activity and lower TIMP-1 level are believed to prolonged and intensify the course of inflammation.

Microbial Population Diversity of the Mud Flat in Suncheon Bay Based on 16S rDNA Sequences and Extracellular Enzyme Activities (남해안 갯벌 미생물의 세포외효소 활성 및 16S rDNA 분석에 의한 다양성 조사)

  • Kim, Yu-Jeong;Kim, Sung-Kyum;Kwon, Eun-Ju;Baik, Keun-Sik;Kim, Jung-Ho;Kim, Hoon
    • Applied Biological Chemistry
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    • v.50 no.4
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    • pp.268-275
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    • 2007
  • Diversity of the mud flat microbial population in Suncheon Bay was investigated by studying extracellular enzyme activities and 16S rDNA sequences. Four culturable bacterial strains with CMCase, xylanase and protease activities were isolated from the wetland and the mud flat. All the strains produced more xylanase activity than CMCase or protease activity, and the properties of the isolate enzymes from the wetland were similar to those from the mud flat. About 2,000 clones were obtained with the 16S rDNA amplified from the metagenomic DNA isolated from the mud samples. Based on the restriction pattern(s), seventeen clones were selected for base sequence analysis. Of the 17 clones, only 35% (6 clones) were found to be cultured strains and 65% (11 clones) to be uncultured strains. The similarities in the base sequences of the clones ranged from 91.0% to 99.9% with an average similarity of 97.3%. The clones could be divided into 7 groups, Proteobacteria (9 clones, 52.9%), Firmicutes (3 clones, 17.6%), Bacteroidetes (1 clone), Flavobacteria (1 clone), Verrucomicrobia (1 clone), Acidobacteria (1 clone), and Chloroflexi (1 clone). Most of the Proteobacteria clones were gamma Proteobacteria associated with oxidation-reduction of sulfur.

Characteristics of Intestinal Microflora and Their Extracellular Pretense of Sardine, Sardinops melanosticta (정어리 내장세균의 특성과 균체외 단백분해효소에 관한 연구)

  • CHANG Dong-Suck;CHO Hak-Rae;CHOI Seung-Tae
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.17 no.3
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    • pp.184-190
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    • 1984
  • Sardine, Sardinops melanosticta, has been caught more than fifty thousand metric tons every year in adjacent sea of Korea, but most of them used for uneatable fish meal because of their rapid spoilage. Usually it is known that the main reason of putrefaction of foods is caused by the maicro-organisms included in them. Therefore, this experiment was carried out to identify the micro-organisms isolated from the intestine of fresh sardine and characterize their proteolytic enzymes produced from them. Aerobic cell count ranged from $1.7{\times}10^4\;to\;3.6{\times}10^5/g$, while anaerobic cell count, from $2.9{\times}10^4\;to\;5.5{\times}10^5/g$. Most of the isolated strains were psychrotrophic mesophiles. Among the two hundred and eighty strains isolated from the fresh samples, fifty-six strains ($20.0\%$) were proteolytics, one hundred and seventy-five strains ($62.5\%$) were lipolytics and tenty-nine strains ($10.5\%$) had the ability to produce hydrogn sulfide. The most predominantly isolated microbial groups from the fresh sardine were Moraxella ($31.4\%$) and Pseudomonas sup. ($28.6\%$). Flavobacterium-Cytophaga, Vibrio, Acinetobacter, Micrococcus spp. and Enterobacteriaceae appeared from $7.9\%\;to\;5\%$ out of total tested strains. The average bacterial count in the spoiled samples (stored at about $18^{\circ}C$ for 48 hours) was increased to the level of $2.9{\times}10^8/g$ for aerobes, $1.5{\times}10^8/g$ for anaerobes, then one hundred and ten strains, corresponding to $52\%$, out of two hundred and thirteen strains submitted to the test were proteolytics. The strongest proteolytic bacterium among the two hundred and eighty strains was identified as Pseudomonas 101 which grew best at $25^{\circ}C$. The optimum condition for the activity of the proteolytic enzyme produced by Pseudomonas 101 appeared $35^{\circ}C$ and pH 9.0, but the activity was relatively unchanged between 5.0 and 11.0 of pH and between $30^{\circ}C\;and\;50^{\circ}C$ of temperature.

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Elimination and Utilization of Pollutants - Part I Microbiological Clarification of Industrial Waste and Its Utilization as Feed Resources - (환경오염원(環境汚染源)의 제거(除去)와 그 이용성(利用性)에 관(關)한 연구(硏究) - 제(報I)1보(第). 미생물(微生物)에 의(依)한 산업폐수(産業廢水)의 정화(淨化) 및 사료자원개발(飼料資源開發)에 개(開)하여 -)

  • Lee, Ke-Ho;Lee, Kang-Heup;Park, Sung-O
    • Applied Biological Chemistry
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    • v.23 no.1
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    • pp.64-72
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    • 1980
  • Industrial wastes from pulp and food plants were treated with microorganisms to clarify organic waste-water and to produce cells as animal feed, and results were summarized as follows. (1) Waste-water from pulp, beer, bread yeast, and ethanol distillation plants contained $1.4{\sim}1.5%$ of total sugar, $0.25{\sim}0.35%$ nitrogen, and biological oxygen demand (BOD) was $400{\sim}25,000$, chemical oxygen demand (COD), $500{\sim}28,000$, and pH, $3.8{\sim}7.0$. The BOD and COD were highest in waste-water from ethanol distillation plants among others. (2) Bacterial and yeast counts were $4{\times}10^4-1{\times}10^9,\;2{\times}10^2-7{\times}10^4/ml$ in waste-water. (3) Bacteria grew better in pulp waste and yeasts in beer, bread yeast, and ethanol distillation waste. (4) Saccharomyces cerevisiae SAFM 1008 and Candida curvata SAFM 70 were the most suitable microorganisms for clarification of ethanol distillation waste. (5) When liquid and solid waste from ethanol distillation were treated with microbial cellulase, xylanase, and pectinase, solid waste was reduced by 36%, soluble waste was increased, and recuding sugar content was increased by 1.3 times which provided better medium than untreated waste for cultivation of yeasts. (6) Optimum growth conditions of the two species of yeast in ethanol distillation waste were pH 5.0, $30^{\circ}C$, and addition of 0.2% of urea, 0.1% of $KH_2PO_4$ and 0.02% of $MgSO_4$. (7) Minimum number of yeast for proper propagation was $1.8{\times}10^5/ml$. (8) C. curvata70 was better than cerevisae for the production of yeast cells from ethanol distillation waste treated with microbial enzymes. (9) S. cerevisiae produced 16 g of dried cell per 1,000ml of ethanol distillation waste and reduced BOD by 46%. C. curvata produced 17.6g of dried cell and reduced BOD by 52% at the same condition. (10) Yeast cells produced from the ethanol distillation waste contained 46-52% protein indicating suitability as a protein source for animal feed.

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Anti-inflammatory Activity of Antimicrobial Peptide Papiliocin 3 Derived from the Swallowtail Butterfly, Papilio xuthus (호랑나비 유래 항균 펩타이드 파필리오신 3의 항염증 활성)

  • Shin, Yong Pyo;Lee, Joon Ha;Kim, In-Woo;Seo, Minchul;Kim, Mi-Ae;Lee, Hwa Jeong;Baek, Minhee;Kim, Seong Hyun;Hwang, Jae Sam
    • Journal of Life Science
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    • v.30 no.10
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    • pp.886-895
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    • 2020
  • The development of novel peptide antibiotics with potent antimicrobial activity and anti-inflammatory activity is urgently needed. In a previous work, we performed an in-silico analysis of the Papilio xuthus transcriptome to identify putative antimicrobial peptides and identified several candidates. In this study, we investigated the antibacterial and anti-inflammatory activities of papiliocin 3, which was selected bioinformatically based on its physicochemical properties against bacteria and mouse macrophage Raw264.7 cells. Papiliocin 3 showed antibacterial activities against E. coli and S. aureus without inducing hemolysis and decreased the nitric oxide production of the lipopolysaccharide-induced Raw264.7 cells. Moreover, ELISA and Western blot analysis revealed that papiliocin 3 reduced the expression levels of pro-inflammatory enzymes, such as inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and prostaglandin E2 (PGE2). In addition, we examined whether papiliocin 3 could inhibit the expression of pro-inflammatory cytokines (interleukin-6 and interleukin-1β) in LPS-induced Raw264.7 cells. We found that papiliocin 3 markedly reduced the expression level of cytokines through the regulation of mitogen-activated protein kinases (MAPK) and nuclear factor kappa B (NF-κB) signaling. We also confirmed that papiliocin 3 binds to bacterial cell membranes via a specific interaction with lipopolysaccharides. Collectively, these findings suggest that papiliocin 3 could be a promising molecule for development as a novel peptide antibiotic.

The Role of Oxygen Free Radicals from Endothelial Cells in Endotoxin-induced Endothelial Cell Cytotoxity (내독소에 의한 혈관 내피세포 손상에서 혈관 내피세포로부터 유리된 산소기의 역할에 관한 연구)

  • Choi, Hyung-Seok;Jeong, Ki-Ho;Yoo, Chul-Gyu;Kim, Young-Whan;Han, Sung-Koo;Shim, Young-Soo;Kim, Keun-Youl;Han, Yong-Chol;Jung, Ki-Suck
    • Tuberculosis and Respiratory Diseases
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    • v.41 no.4
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    • pp.319-327
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    • 1994
  • Background: The pathogenetic mechanism of adult respiratory distress syndrome(ARDS) is not clearly defined yet, but it is well known that increased pulmonary capillary permeabilty is characteristic feature of ARDS. The increased alveolar-capillary permeability is usually preceded by damage of pulmonary artery endothelial cells. The released enzymes and oxygen free radicals from the activated neutrophils seem to play a predominant role in endothelial cell cytotoxicity. The activated neutrophils, however, probably are not the sole contributing factor in this type of damage because many cases of ARDS have been reported in severe neutropenia. Bacterial endotoxin perse and/or oxygen free radicals released from endothelial cells are suggested to be possible factors that contribute to the development of ARDS. The purpose of this study is to investigate the direct cytotoxicity of endotoxin and the role of oxygen free radicals released from the endothelial cells in endotoxin-induced endothelial cell cytotoxicity. Methods: First, to investigate whether endotoxin is cytotoxic to HUVE by itself, various doses of endotoxin were added to culture medium and cytotoxicity was measured. Second, to evaluate the possible role of oxygen free radical in endotoxin-induced HUVE cytotoxicity, various antioxidants were added on the endotoxin-induced HUVE cytotoxicity and cytotoxicity was measured. Third, to verify the release of oxygen free radicals from HUVE, the concentrations of hydrogen peroxide in the endotoxin-treated culture supernatant were measured. Finally, to observe the cytotoxic effect of hydrogen peroxide, HUVE cytotoxicity in the presence of various doses of hydrogen peroxide was measured. The fourth generations of subcultured HUVE from primary culture were used. The cell cytotoxicity was quantified by the chromium-51 release assay. Results: 1) Endotoxin alone showed HUVE cytotoxicity in a dose-dependent fashion. 2) Endotoxin-induced HUVE cytotoxicity was significantly attenuated by the pretreatment of catalase and DMTU. 3) Hydrogen peroxide was released from HUVE after endotoxin treatment in a dose-dependent fashion. 4) Exogenous hydrogen peroxide also showed HUVE cytotoxicity in a dose-dependent fashion. Conclusion: These results suggest that endotoxin alone can directly injure HUVE, and, oxygen-free radicals released from HUVE in response to endotoxin may also participate in the endotoxin-induced HUVE cytotoxicity.

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