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Characterization of Exolytic GH50A β-Agarase and GH117A α-NABH Involved in Agarose Saccharification of Cellvibrio sp. KY-GH-1 and Possible Application to Mass Production of NA2 and L-AHG (Cellvibrio sp. KY-GH-1의 아가로오스 당화 관련 엑소형 GH50A β-아가레이즈와 GH117A α-NABH의 특성 및 NA2와 L-AHG 양산에의 적용 가능성)

  • Jang, Won Young;Lee, Hee Kyoung;Kim, Young Ho
    • Journal of Life Science
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    • v.31 no.3
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    • pp.356-365
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    • 2021
  • Recently, we sequenced the entire genome of a freshwater agar-degrading bacterium Cellvibrio sp. KY-GH-1 (KCTC13629BP) to explore genetic information encoding agarases that hydrolyze agarose into monomers 3,6-anhydro-L-galactose (L-AHG) and D-galactose. The KY-GH-1 strain appeared to possess nine β-agarase genes and two α-neoagarobiose hydrolase (α-NABH) genes in a 77-kb agarase gene cluster. Based on these genetic information, the KY-GH-1 strain-caused agarose degradation into L-AHG and D-galactose was predicted to be initiated by both endolytic GH16 and GH86 β-agarases to generate NAOS (NA4/NA6/NA8), and further processed by exolytic GH50 β-agarases to generate NA2, and then terminated by GH117 α-NABHs which degrade NA2 into L-AHG and D-galactose. More recently, by employing E. coli expression system with pET-30a vector we obtained three recombinant His-tagged GH50 family β-agarases (GH50A, GH50B, and GH50C) derived from Cellvibrio sp. KY-GH-1 to compare their enzymatic properties. GH50A β-agarase turned out to have the highest exolytic β-agarase activity among the three GH50 isozymes, catalyzing efficient NA2 production from the substrate (agarose, NAOS or AOS). Additionally, we determined that GH117A α-NABH, but not GH117B α-NABH, could potently degrade NA2 into L-AHG and D-galactose. Sequentially, we examined the enzymatic characteristics of GH50A β-agarase and GH117A α-NABH, and assessed their efficiency for NA2 production from agarose and for production of L-AHG and D-galactose from NA2, respectively. In this review, we describe the benefits of recombinant GH50A β-agarase and GH117A α-NABH originated from Cellvibrio sp. KY-GH-1, which may be useful for the enzymatic hydrolysis of agarose for mass production of L-AHG and D-galactose.

A comparison study of hygiene status in meals for poorly-fed children through microbiological analysis (결식아동이 이용하는 도시락의 미생물 검사를 통한 위생 상태 비교.분석)

  • Yu, Ok-Kyeong;Kim, Hyun-Suk;Byun, Moon-Sun;Kim, Mina;Cha, Youn-Soo
    • Journal of Nutrition and Health
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    • v.47 no.3
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    • pp.214-220
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    • 2014
  • Purpose: The purpose of this study was to assess hygiene status of meals for poorly-fed children through microbiological quality. Methods: Meals were provided by two social enterprises, one franchise, and one convenience store. There were a total of six meal samples; two samples (social enterprise meal 1; SEM 1, social enterprise meal 2; SEM 2) from two social enterprises, respectively, two samples (franchise meal 1; FM 1, franchise meal 2; FM 2) from one franchise, and two samples (convenience store meal 1; CSM 1, convenience store meal 2; CSM 2) from one convenience store. Microbiological analysis and assessment were performed by Korean food standards codex (KFSC). Results: General bacteria and E. coli in SEM 1 were detected, but the levels were not over KFSC, and Coliform less than $9.2{\times}10$ CFU/g was also detected in seasoned bean sprouts of SEM 1. General bacteria was detected at $1.6{\times}10^6$ CFU/g in cabbage kimchi of SEM 2. Coliform was detected in cabbage kimchi, squid cutlet, stir-fried pork, and fried chicken of FM1 and 2, but the levels were not over KFSC. In addition, S. aureus was detected in cabbage kimchi and seasoned dried white radish of FM 1 and 2 ($9.8{\times}10^2$ CFU/g, $9.4{\times}10^3$ CFU/g respectively), thus was over KFSC. B. cereus was detected in stir-fried pork and fried chicken ($1.2{\times}10^3$ CFU/g, $1.5{\times}10^3$ CFU/g respectively) of FM 1 and 2, thus was over KFSC. Finally, S. aureus was detected in stir-fried dried squid, seasoned spicy chicken, and stir-fried kimchi of CSM 1 and 2, and was over KFSC too ($9.5{\times}10^4$ CFU/g, $2.4{\times}10^2$ CFU/g, $1.3{\times}10^3$ CFU/g respectively). Conclusion: Results of this study suggest that systemic management of hygiene is necessary to safely providing meals to poorly-fed children.

Effect of Traditional Seasoning on Quality Characteristics of Low Temperature Aging Pork (전통 양념이 저온 숙성 돼지고기의 품질특성에 미치는 영향)

  • Jin, S.K.;Kim, I.S.;Hur, S.J.;Park, K.H.;Lyou, H.J.;Kim, I.J.;Hah, K.H.
    • Journal of Animal Science and Technology
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    • v.47 no.6
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    • pp.1041-1050
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    • 2005
  • This study was conducted to investigate the quality characteristics of seasoned pork prepared by Korean traditional 4 types seasoning such as soy sauce (T1); Kimchi sauce (T2); pickled shrimps sauce (T3) and onion sauce (T4). The samples were seasoned by the proportion of meat to seasonings (1:1). The seasoned samples were aged at $1{\pm}1^{\circ}C$ for 30 days. The results obtained were as follows; pH of T3 was higher (p<0.05) than other treatments during aging periods. Saccharinity and salinity were higher in T1. Except T3, water holding capacity (WHC) have no significantly (p>0.05) different during the aging. Shear force and Volatile basic nitrogen (VBN) were increased in T1 and T4 with aging periods. TBARS have no significantly (p>0.05) different in all treatments at the 1 day of aging, however T1 was significantly higher (p<0.05) compared with other treatments after 10 days of aging. In surface meat color, a* and b* were higher in T2 and lower in T4 with aging periods. In inner meat color, L* was decreased in all treatments with aging periods, however T4 was no significantly (p>0.05) different during aging periods. a* and b* were increased with aging periods in all treatments. Total plate counts was increased at the 10 days of aging and decreased at the end of aging. Escherichia coli of T1, T2 and T3 were decreased with aging periods, however T4 was significantly (p<0.05) increased with aging periods. Lactobacilli spp. of T2 and T3 were higher than other treatments at the beginning of aging. In sensory evaluation, T4 was higher at the beginning of aging, however T1 and T3 were increased sensory evaluation value with aging periods.

Antimicrobial, Antioxidant, and Anti-diabetic Activities of Rodgersia podophylla (도깨비부채의 항균, 항산화 및 항당뇨 활성)

  • Pyo, Su-Jin;Lee, Yun-Jin;Kang, Deok-Gyeong;Son, Ho-Jun;Park, Gwang Hun;Park, Jong-Yi;Sohn, Ho-Yong
    • Journal of Life Science
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    • v.30 no.3
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    • pp.298-303
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    • 2020
  • This study aimed to investigate possible applications of Rodgersia podophylla in the food and cosmetic industry. Ethanol extracts of leaves (RP-L), branches (RP-B), and root (RP-R) were prepared, and their antimicrobial, antioxidant, and anti-diabetic activities were evaluated. The polyphenol content in the RP-R, RP-L, and RP-B extracts was 79.6, 30.4, and 16.9 mg/g, respectively. An antimicrobial activity assay showed that the RP-L and RP-R extracts exhibited strong growth inhibition of pathogenic and food spoilage Gram-positive bacteria. Furthermore, the RP-R extract inhibited the growth of the Gramnegative E. coli and P. vulgaris bacteria. All extracts showed strong scavenging activity for DPPH, ABTS, nitrite, and reducing power determined by A 700 nm. In particular, the RC50s of the RP-R extract for the DPPH anion and ABTS cation were 23.0-29.7 and 15.0-18.2 ㎍/ml, respectively, which are comparable to those of vitamin C (9.8 and 8.0 ㎍/ml, respectively). An activity assay of α-glucosidase and β-amylase suggested a high potential for the RP-R extract as an anti-diabetic agent. Its inhibition levels of α-glucosidase and β-amylase at 0.5 mg/ml were 6.9 and 48.5%, respectively. This is the first report of the antimicrobial and anti-diabetic activities of R. podophylla. Our results suggest that RP-L and RP-R extracts could be developed as novel cosmeceutical and functional food resources.

Microbe and Quality Changes of Ready-to-Eat Lettuce during Storage at Different Temperatures (신선편이 양상추의 온도별 저장 중 미생물과 품질변화)

  • Cho, Sun-Kyung;Kwon, Hye-Soon;Park, Jong-Hyun
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.39 no.12
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    • pp.1867-1872
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    • 2010
  • Microbe and quality changes of vacuum-packaged ready-to-eat lettuce were analyzed. While the vacuumpackaged lettuce after chlorine sanitizer were stored at $5^{\circ}C$, $15^{\circ}C$, and $25^{\circ}C$ for 7 days, viable numbers of total aerobic bacteria (TAB), coliform, E. coli, food-borne pathogens and lactic acids bacteria (LAB) were counted with gas production and sensory evaluation. Before the storage, only TAB of 2 log CFU/g and coliform of 1 log CFU/g were detected and LAB was not detected. TAB, coliform and LAB increased by 1 log CFU/g at $5^{\circ}C$ for 7 days without any detection of the pathogens. Sensory evaluations for off-flavour and crispness dropped to half the best value at 5 day storage. TAB and coliform increased by 3 log CFU/g and 2 log CFU/g, respectively, but LAB grew very actively by 4 log CFU/g under anaerobic environment and only B. cereus were detected after enrichment of the lettuce at $15^{\circ}C$ for 3 days. The evaluations for off-flavour and crispness were half the best value for 3 days. However, TAB and coliform increased by 3 log CFU/g, 1 log CFU/g, and 4 log CFU/g, respectively only at 1 day storage under $25^{\circ}C$. Also B. cereus were detected after enrichment and the sensory evaluation were half the best value within 1 day storage. Therefore, preservation at the lowest temperature possible is required for growth inhibition of the bacteria contaminated in the lettuce. Interestingly, LAB among them grew most actively under the anaerobic condition and the adulteration of lettuce might be closely related with the growth of LAB.

Quality Characteristics of Kochujang Prepared with Paecilomyces japonica from Silkworm (누에 동충하초(Paecilomyces japonica)를 첨가하여 제조한 고추장의 품질특성 변화에 관한 연구)

  • Bang, Hye-Yeol;Park, Moo-Hyun;Kim, Gun-Hee
    • Korean Journal of Food Science and Technology
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    • v.36 no.1
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    • pp.44-49
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    • 2004
  • Changes in quality properties of Kochujang prepared with Paecilomyces japonica powder and extract using different solvents were investigated during 90 days of fermentation at $20^{\circ}C$. Although moisture contents were not significantly different, pH of P. japonica-added Kochujang was lower than that of control group without P. japonica, and decreased with increasing fermentation time. Amino nitrogen content increased up to 60 days of fermentation and decreased slightly after 90 days, with that of P. japonica-added Kochujang showing highest on 30 and 60 days at 179.2 and 282.2 mg%, respectively, higher than control gruup. L, a, and b values decreased in proportion to fermentation period, with P. japonica-added Kochujang, particularly P. japonica powder-added Kochujang, lower than those of control g개up. Sensory evaluation test showed color of control group was 'clear red', whereas that of P. japonica powder-added Kochujang was 'dark reddish brown' and P. japonica extract-added Kochujang was darker than control group; consumer preference for dark color was low, Textures of all samples were 'glossy and smooth', showing high consumer preference. Salt content of P. japonica-added Kochujang was higher than that of control group, with P. japonica extract-added Kochujang higher than that made with powder Hot taste or P. japonica-added Kochujang was weaker, whereas its flavor higher, than control group, with P. japonica powder-added Kochujang showing highest flavor score. Overall preference was higher for P. japonica-added Kochujang than control group, with P. japonica water extract-added Kochujang showing the highest score.

Increased Protein of the Secretory Leukocyte Pretense Inhibitor (SLPI) and the Expression of Growth Factors in NIH3T3 Cells by LPS Stimulation (NIH3T3 세포주에서 LPS자극에 의한 분비백혈구단백분해효소억제제 (SLPI)의 단백질증가와 성장인자들의 발현)

  • Lee, Sang-Hwa;Choi, Baik-Dong;Jeong, Soon-Jeong;Jang, Hyun-Seon;Kim, Byung-Ock;Lim, Do-Seon;Park, Joo-Cheol;Wang, Guan-Lin;Jeong, Moon-Jin
    • Applied Microscopy
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    • v.36 no.3
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    • pp.165-172
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    • 2006
  • Secretory leukocyte protease inhibitor (SLPI) involves tissue protection against the destructive action of neutrophil elastase at the site of inflammation. Several studies on new functions of SLPI have demonstrated that SLPI may play a primary role in innate immunity than protease inhibitor, To identify the function of SLPI by lipopolysaccharide (LPS) stimulation in the embryonic fibroblast (NIH3T3) cells. we studied the expression of SLPI compared to other growth factors involving the LPS treatment. To address this, we performed the reverse transcriptase polymerase chain reaction (RT-PCR) and Western blots for the detection of mRNA and protein expression of the SLPI and some growth factors such as VEGF. bFGF, and PDGF-BB after LPS stimulation. NIH3T3 cells were exposed 100 ng/mL Escherichia coli LPS for 30min, 60min, 90min, 24h, and 48h, respectively. The result of RT-PCR showed that SLPI and VEGF mRNA was expressed strongly in NIH3T3 without related to LPS stimulation. mRNA of bFGF was weakly expressed such as the expression of the control. PDGF mRNA expression gradually increased follows at time course. However, SLPI protein level was increased in lysates and culture medium by LPS stimulation. Phase contrast microscopic and scanning electron microscopic observation showed that the increased cell number and cytoplasmic enlargement of the NIH3T3 cells. Therefore, it suggests that the LPS upregulates SLPI expression in NIH3T3 cells. Moreover, secreted SLPI may stimulate cell proliferation and migration.

Screening of the Antimicrobial and Antitumor Activity of Xanthium strumarium L.Extract (한국산 도꼬마리 추출물로부터 항균.항암물질의 탐색)

  • 김현수;유대식;이인선;김용원;여수환
    • KSBB Journal
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    • v.18 no.1
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    • pp.55-61
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    • 2003
  • To isolate and purify the antimicrobial and antitumor agents in Xanthium strumarium L. hydrothermal extract. The crude extract was extracted in ether or ethylacetate under neutral, acidic, and alkali conditions. The antimicrobial activity of each extract was tested against 16 strains of bacteria, 2 strains of yeast, and 2 strains of fungus. The ether neutral extract (XE-N) exhibited the strongest growth inhibition upon the 8 strains of gram-positive bacteria, 6 strains of gram-negative bacteria and Cryptococcus neoformans. Fluorescein diacetate (FDA) testing of XE-N and XEA-N showed growth inhibition of the 3 strains of E. coli, S. aureus and C. albicans even at 30 ng/mL, with the exception of p. aeruginosa. XE-N-S1 and XE-N-S3 from neutral ether extract (XE-N), XE-N-S3 from the acidic ether extract (XE-A), and XEA-N-S1 from ethylacetate (XEA-N) were purified as antimicrobial and antitumor agents. However all purified compounds decomposed with the exception of XE-N-S1. The results upon the antitumor activities of the crude extract and of its purified compounds, showed that XE-N-S1 had the best antitumor activity against HeLa cells. In terms of antitumor activity against HepG2 cells, XE-N-S1 and XE-N-S3 were superior, and against HT29 cells XE-N and XE-N-Sl were good, against Saos2, NCI H522, NCI H1703, Clone M3 cells XE-N-51 was very good, and against LN CAP cells XE-N-S3 was the best. Comparing of cellular toxicities various extracts and purified compounds with the existing antitumor agents, XE-A, XEA-A and XEA-B had the lowest toxicity, and XE-B had a lower toxicity than etoposide. XE-N-S1 and XE-N-S3 showed higher toxicities than etoposide, and the toxicity of XE-A-S3 was higher than that of etoposide, and lower than that of csplatin.

Analysis of Microbiological Contamination in Cultivation and Distribution Stage of Melon

  • Park, Kyeong-Hun;Yun, Hye-Jeong;Kim, Won-Il;Kang, Jun-Won;Millner, Patricia D.;Micallef, Shirley A.;Kim, Byeong-Seok
    • Korean Journal of Soil Science and Fertilizer
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    • v.46 no.6
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    • pp.615-622
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    • 2013
  • The purpose of this study was to evaluate microbial contamination of melons in Korea. A total of 123 samples including melon fruits, leaves, seeds, soils, and irrigation water were collected from farms and markets to detect total aerobic bacteria, coliform, Escherichia coli, and pathogenic bacteria such as Bacillus cereus, Listeria monocytogenes, Salmonella spp., and Staphylococcus aureus. Samples were collected from Iksan and Nonsan farms to monitor bacterial levels on pre-market melons. The total aerobic and coliform bacteria on melon cultivation were between 0.43 and 6.65 log CFU $g^{-1}$, and 0.67 and 2.91 log CFU $g^{-1}$, respectively. Bacillus cereus, a fecal coliform, was detected in soils and melon leaves from Iksan farm at 2.95, 0.73 log CFU $g^{-1}$, respectively, and in soils from Nonsan farm at 3.16 log CFU $g^{-1}$. Market melon samples were collected to assay bacterial load on melon being sold to consumers. The contamination levels of total aerobic bacteria in agricultural markets, big-box retailers, and traditional markets were 4.82, 3.94, 3.99 log CFU $g^{-1}$, respectively. The numbers of coliform in melon on the markets ranged from 0.09 to 0.49 log CFU $g^{-1}$. Listeria monocytogenes, Salmonella spp., and Staphylococcus aureus were not detected in any samples. The count of total aerobic bacteria on melon seeds ranged from 0.33 to 3.34 log CFU $g^{-1}$. This study found that irrigation water, soil, manure and various farm work activities including post-harvest processes were latent sources of microbial contamination. These results suggest that hygienic management and monitoring of soil, water, and agricultural material should be performed to reduce microbial contamination in melon production.

Effect of SeaR gene on virginiamycins production in Streptomyces virginiae (희소방선균 SeaR 유전자가 Streptomyces virginiae의 virginiamycins 생산에 미치는 영향)

  • Ryu, Jae-Ki;Kim, Hyun-Kyung;Kim, Byung-Won;Kim, Dong-Chan;Lee, Hyeong-Seon
    • Korean Journal of Microbiology
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    • v.51 no.3
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    • pp.256-262
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    • 2015
  • In order to study the effect of the receptor protein (SeaR), which is isolated from Saccharopolyspora erythraea, we introduced the SeaR gene to Streptomyces virginiae as host strains. An effective transformation procedure for S. virginiae was established based on transconjugation by Escherichia coli ET12567/pUZ8002 with a ${\varphi}C31$-derived integration vector, pSET152, which contained int, oriT, attP, and $ermEp^{\ast}$ (erythromycin promotor). Therefore, the pEV615 was introduced into S. virginiae by conjugation and integrated at the attB locus in the chromosome of the recipients by the ${\varphi}C31$ integrase (int) function. Transformants of S. virginiae containing the SeaR gene were confirmed by PCR and transcriptional expression of the SeaR gene in the transformants was analyzed by RT-PCR, respectively. And, we examined the production time of virginiamycins in the culture media of both the transformants and the wild type. The production time of virginiamycins in the wild type and transformants was the same. When 100 ng/ml of synthetic $VB-C_6$ was added to the state of 6 or 8 hour cultivation of wild type and transformants, respectively, the virginiamycins production was induced, meaning that the virginiamycins production in the wild type was detected 2 h early than transformants. From these results, SeaR expression was also affected to virginiamycins production in transformants derived from S. virginiae. In this study, we showed that the SeaR protein worked as a repressor in transformants.