• KSBB Journal
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• 제32권3호
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• pp.193-198
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• 2017

Sialylation is important in producing therapeutic proteins such as antibody, cytokine and fusion protein. Thus, enhancement of sialylation is usually performed in CHO cell cultures. ${\alpha}2,6$-Sialyltransferase (ST), which plays a key role in the attachment of ${\alpha}2,6-sialic$ acid, is present in human cells but not in Chinese hamster ovary (CHO) cells. Overexpression of ${\alpha}2,6-ST$ can be used for enhancing the degree of sialylation and achieving human-like glycosylation. In this study, we constructed CHO cells producing human cytotoxic T-lymphocyte antigen4-immunoglobulin (hCTLA4-Ig) as well as ${\alpha}2,6-ST$. Transfected CHO cells were selected using G418 and stable cell line was established. Profiles of viable cell density and hCTLA4-Ig titer in an overexpressed cell line were similar to those of a wild-type cell line. It was confirmed that the total amount of sialic acid was increased and ${\alpha}2,6-sialic$ acid was attached to the terminal residues of N-glycan of hCTLA4-Ig by ESI-LC-MS. Compared to 100% of ${\alpha}2,3-sialic$ acid in wild type cells, 70.9% of total sialylated N-glycans were composed of ${\alpha}2,6-sialic$ acid in transfected cells. In conclusion, overexpression of ${\alpha}2,6-ST$ in CHO cells led to the increase of both the amount of total sialylated N-glycan and the content of ${\alpha}2,6-sialic$ acid, which is more resemble to human-like structure of glycosylation.

• Molecules and Cells
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• 제37권2호
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• pp.172-177
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• 2014

A glycosyltransferase, YjiC, from Bacillus licheniformis has been used for the modification of the commercially available isoflavonoids genistein, daidzein, biochanin A and formononetin. The in vitro glycosylation reaction, using UDP-${\alpha}$-D-glucose as a donor for the glucose moiety and aforementioned four acceptor molecules, showed the prominent glycosylation at 4' and 7 hydroxyl groups, but not at the $5^{th}$ hydroxyl group of the A-ring, resulting in the production of genistein 4'-O-${\beta}$-D-glucoside, genistein 7-O-${\beta}$-D-glucoside (genistin), genistein 4',7-O-${\beta}$-D-diglucoside, biochanin A-7-O-${\beta}$-D-glucoside (sissotrin), daidzein 4'-O-${\beta}$-D-glucoside, daidzein 7-O-${\beta}$-D-glucoside (daidzin), daidzein 4', 7-O-${\beta}$-D-diglucoside, and formononetin 7-O-${\beta}$-D-glucoside (ononin). The structures of all the products were elucidated using high performance liquid chromatography-photo diode array and high resolution quadrupole time-of-flight electrospray ionization mass spectrometry (HR QTOF-ESI/MS) analysis, and were compared with commercially available standard compounds. Significantly higher bioconversion rates of all four isoflavonoids was observed in both in vitro as well as in vivo bioconversion reactions. The in vivo fermentation of the isoflavonoids by applying engineered E. coli $BL21(DE3)/{\Delta}pgi{\Delta}zwf{\Delta}ushA$ overexpressing phosphoglucomutase (pgm) and glucose 1-phosphate uridyltransferase (galU), along with YjiC, found more than 60% average conversion of $200{\mu}M$ of supplemented isoflavonoids, without any additional UDP-${\alpha}$-D-glucose added in fermentation medium, which could be very beneficial to large scale industrial production of isoflavonoid glucosides.

• 한국미생물·생명공학회지
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• 제24권1호
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• pp.19-26
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• 1996

A phospholipase C (PLC) inhibitor (MT267-2B) was isolated from the culture broth of actinomycetes isolate MT2617-2 by the extraction with n-butanol and column chromatographic techniques. The molecular weight of the inhibitor was 1057, by the spectroscopic analyses of IR, $^{13}C$-and $^{1}H$-NMR and ESI-MS. The chemical structure of MT2617-2B was found to be a macrolide compound consisted of a hemiketal ring, polyhydroxyl and polymethyl groups, which had a malonate and guanidine group as its side chain. MT2617-2B produced its two isomers having the same molecular weight by standing in methanol solution at room temperature. Therefore, MT2617-2B was identified as copiamycin and niphithricin A, macrolide antibiotics. The values of $IC_{50}$ against PLC-${\gamma}$1 and PLC-${\beta}$1 were 25 and 50${\mu}$g/ml, respectively. MT2617-2B had antimicrobial activities against Staphylococcus aureus and Candida albicans, but not against Escherichia coli.

• 한국유기농업학회:학술대회논문집
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• 한국유기농학회 2009년도 하반기 학술대회
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• pp.319-319
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• 2009

관행재배 농산물에 대한 유기재배 농산물의 우수성이 구전되고 있으나 객관적인 과학적 데이터가 부족한 실정이다. 그래서 본 연구에서는 한국인이 주식으로 하고 있는 쌀(동진 1호)을 대상으로 생산이력이 분명한 유기 및 관행재배 시료를 이용하여 독자적으로 확립한 분석법을 이용해 유기 및 관행재배 쌀에 함유된 생리활성성분의 함량 비교를 행하였다. 생리활성성분 분석은 $\gamma$-oryzanol, $\beta$-sitosterol 및 tocopherol류를 대상으로 행하였다. 쌀에 함유된 주요 4종의 $\gamma$-oryzanol류의 동정을 위해 그 4종 화합물들을 ESI-MS 및 NMR 분석을 행한 결과, ODS 컬럼 상에서 cycloartenyl ferulate, 24-methylene cycloartenyl ferulate, campesteryl ferulate 및 $\beta$-sitosteryl ferulate 순으로 용출됨을 확인하였다. 동정된 각 성분들을 HPLC로 분석한 결과, cycloartenyl ferulate와 $\beta$-sitosteryl ferulate의 함량은관행보다 유기재배 쌀에서 유의(p<0.05)하게 높은 값을 보였다. 또한 $\gamma$-oryzanol 중 24-methylene cycloartenyl ferulate와 campesteryl ferulate의 함량은 유의차는 인정되지 않았으나 관행보다 유기재배 쌀에서 더 높은 경향을 보였다. 그리고 $\beta$-sitosterol 함량에 있어서는 유의차가 인정되지 않았으나 관행보다 유기재배 쌀에서 더 높은 경향을 나타냈다. 관행재배와 유기재배 쌀의 tocopherol류의 함량을 분석한 결과, $\alpha$-tocopherol의 함량에 있어서는 관행과 유기재배 쌀 간의 유의차가 관찰되지 않았고, $\beta$- and/or $\gamma$-tocopherols의 함량에 있어서는 유기재배 쌀에서 더 높은 경향이 관찰되었다. 이처럼 쌀에 함유된 생리활성 성분과 그로 인해 발휘되는 기능성 측면에 있어서 유기농산물의 우수성은 유기농산물 재배 농가의 수익 창출에 도움이 되는 홍보자료 제공 및 식품 기능학적 측면에 있어 중요한 기초자료로 활용될 수 있으리라 기대된다. 그러나 본 결과는 본 실험에서 대상으로 한 시료에 한정될 수 있으며, 유기재배 방법과 재배환경 및 조건에 따라 다양한 결과가 얻어질 수 있음에 주의가 요구된다.

• 생약학회지
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• 제49권1호
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• pp.47-54
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• 2018

Astilbe rubra (AR) is a perennial, belongs to the Saxifragaceae family, it contains tannin and triterpene. AR has been used in republic korea to improve toxication, fever, pain and convulsion. Recently, number of natural products have been analyzed for potential pharmacological activities including anti-cancer, anti-obesity and anti-diabetic medication. Consequently, we investigated the growth inhibition effect of Astilbe rubra water extract (WAC), ethanol extract (EAC) and bergenin on Hela cell (human adenocarcinoma cell). From whole plant of A. rubra, bergenin was isolated by column chromatography and its structures were identified by $^1H$, $^{13}C$ NMR and IT TOF-ESI MS. High extraction efficiency of bergenin was shown at 0.95% under 60 min reflux extraction with 50% MeOH. The MTS assay showed that EAC (ethanol extract) treatment increased cell death in a dose-dependent manner. Moreover, EAC treatment on Hela cell increased apoptotic cell death and caspase-3 activity. Results suggest that EAC has growth inhibition effect on Hela cells, but not WAC and bergenin. $500{\mu}g/mL$ EAC treatment inhibited Hela cell at $60.2{\pm}1.5%$.

• 한국식품영양학회지
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• 제26권3호
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• pp.547-552
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• 2013

황백 껍질은 황벽나무(Phellodendron amurense)의 건조된 수피로부터 얻어진다. 이 수피는 한국의 전통 한약제로서, 설사, 황달, 무릎과 발의 통증, 요도관 및 피부 감염증에 폭넓게 사용되어 왔다. 본 연구는 황벽나무의 메탄올 추출액으로부터 항균성 화합물 분리를 위해 CPC 방법으로 효과적으로 수행하였다. 두 용매의 CPC 최적조성은 n-butanol:acetic acid:water(4:1:5 v/v/v)이었다. 이동상의 유속은 1,000 rpm 회전력에서 상승법으로 분당 3 $m{\ell}$ 속도로 전개시켰다. CPC에서 분리된 분획분은 prep-HPLC로 정제하였다. 분리된 palmatine은 $^1H$, $^{13}C$-NMR, ESI-MS 데이터 분석으로 확인하였다.

• 생약학회지
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• 제47권3호
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• pp.211-216
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• 2016

In the course of our continuing search for biologically active components from Korean medicinal plants, we isolated the main compound, luteolin 5-glucoside from aqueous fraction of Ajuga spectabilis. The structure was elucidated by the basis of $^1H$ and $^{13}C$ NMR and TOF ESI-MS data. Quantitative analysis of luteolin 5-glucoside was carried out on a XBridge C18 column ($S-5{\mu}m$, $4.6{\times}250mm$) with gradient elution composed of acetonitrile:water. The results exhibit that the average content of main compound in A. spectabilis were 0.048%. Oxidative stress plays a major role Alzheimer's disease (AD) and other neurodogenerative disease. AD is major health problem and there is currently no clinically accepted treatment to cure or stop its progression. Pretreatment with luteolin 5-glucoside markedly attenuated $H_2O_2$-induced cell viability loss in a dose-dependent manner. Luteolin 5-glucoside also inhibited the formation of intracellular reactive oxygen species in SH-SY5Y. The results suggest that luteolin 5-glucoside from A. spectabilis has protective effects against oxidative stress-induced cytotoxicity, which might be a potential therapeutic compound for treating and/or preventing neurodegenerative disease implicated with oxidative stress.

• 생약학회지
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• 제47권3호
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• pp.232-236
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• 2016

In traditional Korean medicine, Osmunda japonica Thunb has been used as hemostasis and antipyretic treatment. The main compound "cinnamtannin B-1" was obtained by column chromatographic separation, and its structure was determined by spectroscopic methods, including $^1H$, $^{13}C$ NMR, and IT-TOF-ESI MS. Ash, moisture and extract content and acidinsoluble ash were monitored as identification test to establish the analytical methods. The optimum reflux extraction condition was 100% methanol extracted 30 min for 2 times. A quantitative analysis using HPLC method exhibited that the main compound at 24.7 min and its content was 0.96% in methanol extraction.

• 생약학회지
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• 제49권2호
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• pp.138-144
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• 2018

Asparagus oligoclonos is one of the endemic halophytes used folk medicine in Korea. We isolated the main compound rutin from methanol extracts of A. oligoclonos based on nuclear magnetic resonance and TOF ESI-MS data. We have investigated the quantitative analysis method of main compound using HPLC and the results exhibit that rutin content of A.oligoclonos were 1.816%. To explore anti-oxidant from A. oligoclonos ethanol extracts (AOE), we investigated the antioxidant effects of AOE on $H_2O_2$-induced oxidative stress in RAW 264.7 cell. AOE were reduced $H_2O_2$-induced oxidative stress via enhancement of cell viability, and AOE significantly decreased ROS production depending on concentration. Next, to screen for anti-inflammatory activity of AOE, we investigated the inhibitory effects of AOE in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. AOE had no effect on cell viability at a concentration of $500{\mu}g/mL$. Nitric oxide (NO) production inhibited in a dose-dependent manner. These results suggest that AOE may be a useful antioxidant and anti-inflammatory agent.

• 한국식품저장유통학회지
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• 제19권2호
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• pp.229-234
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• 2012

양파의 매운 맛과 냄새를 감소시켜 섭취가 용이하도록 제조된 흑양파가 항산화 활성이 있다고 보고되어있어서 흑양파로부터 항산화 활성물질을 구명하고자 하였다. 흑양파 MeOH 추출물을 용매분획하여 얻어진 분획물들 중 EtOAc층이 높은 DPPH와 $ABTS^+$ radical scavenging 활성이 있었다. 그래서 EtOAc층을 silica gel과 Sephadex LH-20 등의 column chromatography를 이용하여 2종의 항산화 활성물질을 분리하였다. 단리한 이들 화합물을 대상으로 ESI-MS 및 NMR 분석을 통하여 3,4-dihydroxybenzoic acid (1)와 quercetin (2)로 각각 동정하였다. 이 화합물들은 $ABTS^+$ 및 DPPH radical scavenging 활성이 있었으며, 화합물 2는 화합물 1에 비해 더 높은 radical scavenging 활성이 있었다.