• Title/Summary/Keyword: substrate inhibition

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Rare ginsenoside Ia synthesized from F1 by cloning and overexpression of the UDP-glycosyltransferase gene from Bacillus subtilis: synthesis, characterization, and in vitro melanogenesis inhibition activity in BL6B16 cells

  • Wang, Dan-Dan;Jin, Yan;Wang, Chao;Kim, Yeon-Ju;Perez, Zuly Elizabeth Jimenez;Baek, Nam In;Mathiyalagan, Ramya;Markus, Josua;Yang, Deok-Chun
    • Journal of Ginseng Research
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    • v.42 no.1
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    • pp.42-49
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    • 2018
  • Background: Ginsenoside F1 has been described to possess skin-whitening effects on humans. We aimed to synthesize a new ginsenoside derivative from F1 and investigate its cytotoxicity and melanogenesis inhibitory activity in B16BL6 cells using recombinant glycosyltransferase enzyme. Glycosylation has the advantage of synthesizing rare chemical compounds from common compounds with great ease. Methods: UDP-glycosyltransferase (BSGT1) gene from Bacillus subtilis was selected for cloning. The recombinant glycosyltransferase enzyme was purified, characterized, and utilized to enzymatically transform F1 into its derivative. The new product was characterized by NMR techniques and evaluated by MTT, melanin count, and tyrosinase inhibition assay. Results: The new derivative was identified as (20S)-$3{\beta},6{\alpha},12{\beta}$,20-tetrahydroxydammar-24-ene-20-O-${\beta}$-D-glucopyranosyl-3-O-${\beta}$-D-glucopyranoside(ginsenoside Ia), which possesses an additional glucose linked into the C-3 position of substrate F1. Ia had been previously reported; however, no in vitro biological activity was further examined. This study focused on the mass production of arduous ginsenoside Ia from accessible F1 and its inhibitory effect of melanogenesis in B16BL6 cells. Ia showed greater inhibition of melanin and tyrosinase at $100{\mu}mol/L$ than F1 and arbutin. These results suggested that Ia decreased cellular melanin synthesis in B16BL6 cells through downregulation of tyrosinase activity. Conclusion: To our knowledge, this is the first study to report on the mass production of rare ginsenoside Ia from F1 using recombinant UDP-glycosyltransferase isolated from B. subtillis and its superior melanogenesis inhibitory activity in B16BL6 cells as compared to its precursor. In brief, ginsenoside Ia can be applied for further study in cosmetics.

Anti-bacterial Effects of Aqueous Extract Purified from the Immature Cone of Red Pine (Pinus densiflora) (미성숙 솔방울 열수추출물의 항균성)

  • Jeong, Kyung Hui;Hwang, In Sik;Kim, Ji Eun;Lee, Young Ju;Kwak, Moon Hwa;Lee, Young Hee;Lee, Jae Ho;Hwang, Dae Youn;Jung, Young Jin
    • Textile Coloration and Finishing
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    • v.26 no.1
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    • pp.45-52
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    • 2014
  • Red pine (Pinus densiflora) is widely used traditional medicine, pharmacological and nutritional values from which the phytochemical compounds are derived. The present study was aimed to examine the antibacterial effects in the absence and presence of a immature red pine cone extract against 13 microorganisms. The components in the aqueous extract from immature red pine cone were identified by GC-MS. About 1.4% of total polyphenolic compound was measured in aqueous extract collected from immature red pine cone. Also, the high concentration of ${\beta}$-phellenandrene, ${\alpha}$-pinene, limonene, bornyl acetate and aldehyde was detected in total ion chromatograms. Of total 13 microorganisms, 4 microorganisms including Pseudomonas aeruginosa, Vibrio cholera, Listeria monocytogenes, Klebsiella pneumonia were effectively killed by aqueous extract of immature red pine cone. The highest anti-bacterial effect was detected in P. aeruginosa, followed by V. cholera, L. monocytogenes and K. pneumonia. In case of P. aeruginosa, the largest diameter of inhibition zone was maintained to 1/2 solution treated cells and slightly decreased at 1/4 and 1/8 solution treated cells. Also, in test used V. cholera and L. monocytogenes, the inhibition zone was strongly formed in only 1 and 1/2 solution treated cells, while K. pneumonia showed the very small diameter of inhibition zone in all concentrations. Therefore, these results suggested that the aqueous extracts of immature red pine cone should be considered as a new and potentially important anti-bacterial substrate to effectively prevent the microbial infection and penetration.

Antimicrobial and Antioxidative Activities of Solvent Fraction from Humulus japonicus (환삼덩굴의 용매분획별 항균성 및 항산화성)

  • Park, Seung-Woo;Woo, Cheol-Joo;Chung, Shin-Kyo;Chung, Ki-Taek
    • Korean Journal of Food Science and Technology
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    • v.26 no.4
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    • pp.464-470
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    • 1994
  • The biological activities of Humulus japonicus were extracted by water and methanol. Methanol was better solvent than water in the extraction for antimicrobial activities against six different species of bacteria and two yeasts. The methanol extract was systematically fractionated with various organic solvents which have different polarities. From the result of antimicrobial activities against six species of bacteria and two species of yeasts, methanol extract was superior to water extract. The methanol extract of Humulus japonicus showed antimicrobial activity against the all species of microorganisms tested except Escherichia coli . The butanol fraction of methanol extract showed antimicrobial effect on the all species tested. The minimal inhibition concentration(MIC) of the butanol fraction on the growth of microorganisms was ranged between $0.1{\sim}0.4%$. The water extract of Humulus japonicus did not show inhibition of the activity of trypsin but methanol extract showed inhibitory activity. The chloroform fraction of methanol extract showed comparatively higher trypsin inhibitory activity than other fractions. The concentration of 50% inhibition$(IC_{50})$ by chloroform fraction was 1.0 mg/ml. Enzyme-inhibitor complex formation was above 90% of the while for 20 min. It was revealed that methanol extract of Humulus japonicus inhibited peroxide production of lard and soybean oil as substrate by antioxidative test. The chloroform fraction of methanol extract had the highest activity. When 0.2% of chloroform fraction was added, induction period of soybean oil and lard were extended 15, 9 days, respectively.

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Comparative molecular field analysis (CoMFA) and holographic quantitative structure-activity relationship (HQSAR) on the growth inhibition activity of the herbicidal 3-phenyl-5-(3,7-dichloro-8-quinolinyl)-1,2,4-oxadiazole derivatives (제초성 3-Phenyl-5-(3,7-dichloro-8-quinolinyl)-1,2,4-oxadiazole 유도체들의 생장 저해활성에 관한 비교 분자장 분석 (CoMFA)과 분자 홀로그램 구조-활성관계 (HQSAR))

  • Sung, Nack-Do;Lee, Sang-Ho;Song, Jong-Hwan;Kim, Hyoung-Rae
    • The Korean Journal of Pesticide Science
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    • v.7 no.2
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    • pp.108-116
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    • 2003
  • A series of new quinclorac family, herbicidal 3-phenyl-5-(3,7-dichloro-8-quinolinyl)-1,2,4-oxadiazole derivatives as substrate were synthesized and their growth inhibition activity $(pI_{50})$ against root and shoot of rice plant (Oryza sativa L.) and barnyard grass (Echinochloa crus-galli) were determined. And then comparative molecular field analysis (CoMFA) and molecular holographic quantitative structure- activity relationship (HQSAR) were compared in terms of their potential for predictiability. The statistical results were suggested that HQSAR based model had better predictability than CoMFA model. The selective factors to remove barnyard grass take electron withdrawing groups which can be created positive charge and steric bulky on the phenyl ring. Results revealed that the unknown 2,6-dichloro-substituent, U5 and 2,4,6-trichloro-substituent, U6(${\Delta}pI_{50}$=CoMFA: 1.18 & HQSAR: 1.82) were predicted as compound with higher activity and selectivity.

Induction of Apoptosis in Human Colon Carcinoma HCT116 Cells Using a Water Extract of Lepidium virginicum L. (콩다닥냉이 추출물에 의한 HCT116 대장암세포의 사멸 유도에 관한 연구)

  • Chae, Yang-Hui;Shin, Dong-Yeok;Park, Cheol;Lee, Yong-Tae;Moon, Sung-Gi;Choi, Yung-Hyun
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.40 no.5
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    • pp.649-659
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    • 2011
  • To examine the anti-cancer effects of Lepidium virginicum L., the anti-proliferative and pro-apoptotic effects of a water extract of L. virginicum leaves (WELVL) and of L. virginicum roots (WELVR) were investigated in HCT116 human colon carcinoma cells. The treatment of HCT116 cells with WELVL and WELVR resulted in the inhibition of growth and morphological changes in a concentration-dependent manner by inducing apoptosis. The growth inhibition and apoptosis induction by WELVR was stronger than that of WELVL thus, we determined that WELVR was the more optimal extract for this study. The increased apoptotic events in HCT116 cells caused by WELVR were associated with an up-regulation of Fas ligand, Bax, and Bad expression, a down-regulation of Bcl-2, Bcl-$_XL$, and Bid expression, and a decrease in the mitochondrial membrane potential (MMP, ${\Delta}{\psi}m$). WELVR treatment induced the proteolytic activation of caspase-3, -8, and -9, and the degradation of caspase-3 substrate proteins, such as poly (ADP-ribose) polymerase (PARP), ${\beta}$-catenin, and phospholipase C-${\gamma}1$ (PLC-${\gamma}1$). In addition, apoptotic cell death induced by WELVR was correlated with a down-regulation of inhibitors of the apoptosis protein (IAP) family, such as the X-linked inhibitor of apoptosis protein (XIAP), cIAP-1, and cIAP-2. These findings suggest that the WELVR-induced inhibition of cell proliferation is associated with the induction of apoptotic cell death. WELVR may be a potential chemotherapeutic agent for the control of HCT116 human colon carcinoma cells.

Hydrolysis of Fish Protein Concentration in an Ultrafiltration Membrane Reactor (한외여과막 반응기를 이용한 FPC의 가수분해)

  • 최정호;변희국;김세권
    • Membrane Journal
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    • v.10 no.2
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    • pp.83-91
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    • 2000
  • In order to improve functional properties, enzymatic hydrolysis of FPC (fish protein concentration) was achieved in ultrafiltration membrane reactor (MWCO 5,000). First, insoluble FPC was hydrolyzed by pepsin in batch reactor to decrease the fouling in ultrafiltration membrane reactor, and second hydrolysis was achieved by pronase E in ultrafiltration membrane reactor The optimum operating conditions in batch reactor using pepsin were at temperature 45$^{\circ}C$, pH 2.0 and the ratio of substrate to pepsin, 150 (w/w) After operating for 5hrs under optimum conditions, 89% of total amount of initial FPC was hydrolyzed. The rate constants, $K_{m}$ and V$_{max}$, were 1.25% and 0.89 mg/$m\ell$/min, respectively, and substrate inhibition was occured above 1.5%. The ultrafiltration membrane reactor was operated with recycling rate of 474 $m\ell$/min and transmembrane pressure of 15 psi. The permeate flux was increased by temperature, transmembrane pressure, but the permeate flux was fixed by pH. The optimum ratio of substrate to pronase E was 200(w/w) and the productivity of ultrafiltration membarane reactor was 702 mg/mg -enzyme, that of batch reactor was 51mg/mg-enzyme. Molecular weight distributions tot first and second hydrolysates were from 2,500 Da to 20,000 Da and from 700 Da to 10,000 Da, respectivelyly.

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Effect of chloride ions on the catalytic properties of human pancreatic α-amylase isozyme produced in Pichia pastoris (Pichia pastoris에서 생산된 인체 췌장 α-아밀레이스 동질효소의 촉매활성에 대한 염소이온의 영향)

  • Kim, Min-Gyu;Kim, Young-Wan
    • Korean Journal of Food Science and Technology
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    • v.48 no.4
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    • pp.341-346
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    • 2016
  • The AMY2B gene, encoding human pancreatic ${\alpha}$-amylase isozyme (HPA II), was expressed in Pichia pastoris, and the effects of chloride ions on HPA II activity toward starch substrates were investigated. As seen with chloride ion-dependent ${\alpha}$-amylases-including HPA I, the isozyme of HPA II-chloride ions increased enzyme activity and shifted the optimal pH to an alkaline pH. The activity enhancement by chloride was more significant at pH 8 than that at pH 6, suggesting that the protonation state of the general acid/base catalyst of HPA II was important for the hydrolysis of starches at an alkaline pH because of the increase in its $pK_a$ by chloride ions. The turnover values for cereal starches as the substrates markedly increased in the presence of chloride by up to 7.2-fold, whereas that for soluble starch increased by only 1.7-fold. Chloride inhibited substrate hydrolysis at high substrate concentrations, with $K_i$ values ranging from 6 to 15 mg/mL.

Hydrolysis of 7S and 11S Soy Proteins by Commercial Proteases (단백분해효소(蛋白分解酵素)에 의한 대두(大豆) 7S 및 11S 단백질(蛋白質)의 가수분해(加水分解))

  • Kang, Yeung-Joo;Lee, Ki-Chun;Park, Yeung-Ho
    • Korean Journal of Food Science and Technology
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    • v.20 no.3
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    • pp.338-343
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    • 1988
  • Selected kinetic parameters and degree of hydrolysis(DH) were measured using commercial proteases(trypsin, alcalase and pronase) to study the affinity of these enzymes to 7S and 11S soy proteins. Electrophoretic patterns of the hydrolysates were also investigated. In general, the order of affinity between the proteins and the proteases was 11S(protein-rich fraction)and 7S PRF for unheated proteins, and 7S PRF and 11S PRF for preheated proteins. Substrate inhibition was present at a substrate concentration of 1.5% or higher when preheated protein was used as the substrate. The maximum DH values of alcalase were obtained from 7S PRF(60%) and 11S PRF(80%) at 1 hr hydrolysis, respectively. Trypsin hydrolyses did not affect 11S soy protein but the acidic subunits in contrast to alcalase and pronase hydrolyses which changed almost all subunits. Alcalase hydrolysis induced distinct changes on 2S soy protein.

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Curing Properties of UV-LED Curable Color Coating (UV-LED 경화형 칼라 코팅의 경화특성 연구)

  • Ho, Shin-Chan;Kim, Jong-Gu;Hong, Jin-Who;Ahn, Tae-Jung;Kim, Hyun-Kyoung
    • Journal of Adhesion and Interface
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    • v.13 no.1
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    • pp.31-37
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    • 2012
  • In this study, we investigated the curing properties of UV-LED curable color coating. Specially, the effects of UV-LED wavelength (365, 395, and 405 nm), inert gas, the concentration of photosensitizer, and dual curing on the curing behavior of UV-LED coating systems were studied. The photopolymerization behaviors and the unreacted acrylate groups at Film-air (FA) interface and Film-substrate (FS) interface were investigated by photo differential scanning calorimetry (Photo-DSC) and Fourier-transform infrared spectroscopy with attenuated total reflection (FT-IR/ATR), respectively. Photo-DSC results showed that the heat flow and the ultimate conversion for coating system cured by 405 nm UV-LED were higher than the corresponding values for coating systems cured by 395 and 365 nm UV-LED. FT-IR/ATR results showed that the UV-LED curing systems improved interior and through curing of the coating film, but significantly are affected by oxygen inhibition at FA-interface. The inert environment such as nitrogen purging and the dual curing improved the surface and interior curing of the coating films.

Synergism among Endo-xylanase, $\beta$-Xylosidase, and Acetyl Xylan Esterase from Bacillus stearothermophilus

  • Suh, Jung-Han;Choi, Yong-Jin
    • Journal of Microbiology and Biotechnology
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    • v.6 no.3
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    • pp.173-178
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    • 1996
  • Synergic effects among endo-xylanase, $\beta$-xylosidase, and acetyl xylan esterase of Bacillus stearothermophilus in the hydrolysis of xylan were studied by using birchwood, oat spelt, and acetylated xylan as substrates. Synergism between endo-xylanase and $\beta$-xylosidase was observed on all three substrates tested, indicating that $\beta$-xylosidase enhanced the production of xylose by relieving the end-product inhibition upon endo-xylanase conferred by xylooligomers. Endo-xylanase and $\beta$-xylosidase also showed synergism with acetyl xylan esterase in the hydrolysis of birchwood and acetylated xylan, while no synergic effect was detected in oat spelt xylan hydrolysis. Thus, the hydrolysis of xylan containing acetic acid side chains required the action of acetyl xylan esterase, which eliminated the steric hindrance of the side chains, leading to the better hydrolysis by endo-xylanase and $\beta$-xylosidase , and the acetyl xylan esterase activity was also enhanced by endo-xylanase and $\beta$-xylosidase for the latter enzymes provided acetyl xylan esterase with shorter xylan oligomers, the better substrate for the enzyme.

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