• Title/Summary/Keyword: Functional assay

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Functional Expression of a Dipeptide Transporter Obtained from Intestinal HT-29 Cells Using Xenopus Oocytes (장관세포인 HT-29에 존재하는 디펩티드수송체의 Xenopus oocyte에서의 발현)

  • Oh, Doo-Man;Yang, Chae-Ha
    • Journal of Pharmaceutical Investigation
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    • v.25 no.4
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    • pp.299-305
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    • 1995
  • Cloning the gene encoding a dipeptide transporter is necessary for understanding the absorption mechanism of peptides and peptide-like drugs in the gastrointestinal tract. Functional expression of a dipeptide transporter after microinjection into Xenopus laevis oocytes was performed using the mRNA purified from human intestinal HT-29 cells. Fifty nanoliters of purified mRNA (1 mg/mL) were microinjected into healthy oocytes followed by incubation for 4 days in order to express a dipeptide transporter. Functional expression was determined by a uptake assay using 10 Ci/mL $[^3H]-glycylsarcosine$, a dipeptide substate of the transporter. Seasonal variability and batch-to-batch variability were greater in summer. The usage of beveled micropipettes improves viability of oocytes at 4 days after microinjection. Expression of a dipeptide transporter in oocytes after microinjection of mRNA obtained from HT-29 cells was significantly larger than those after microinjection of water or mRNA obtained from the rabbit intestine.

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Development of Microwave Extraction Method for the Active Ingredients and Functional Constituents of Paeonia Root (마이크로웨이브 추출방법을 이용한 작약의 유효성분 추출 및 생리활성 측정)

  • Lee, Hyojin;Jang, Kyoung won
    • Korean Journal of Pharmacognosy
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    • v.52 no.3
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    • pp.157-162
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    • 2021
  • The heat-mediated reflux apparatus extraction of Paeonia lactiflora Pall. has been widely used as a traditional extraction method. In this paper, the microwave apparatus extraction method of Paeonia Radix was performed and the active ingredients and functional constituents were compared with the reference extraction method. The most effective extraction condition of albiflorin using the microwave was 120℃ in 50% methanol, and paeoniflorin was maximally extracted at 60℃. The reduced level of paeoniflorin molecule at high-pressure and high-temperature extraction condition was caused by the molecular instability. Additionally, the microwave extraction of 50% methanol extracts at 150℃ showed the highest functional constituents determined by in vitro DPPH radical scavenging activity, polyphenol concentration, and tyrosinase inhibition assay. The microwave apparatus was adapted as a rapid, low-cost, and environmentally friendly method to extract active ingredients and the practical extraction conditions of Paeonia Radix can be used in industrial applications.

Development of Selectable Marker of High Oleate Trait in Peanut (Arachis hypogaea L.) (땅콩에서 고 올레인산 형질관련 분자마커의 선발)

  • Yang, Kiwoung;Pae, Suk-Bok;Park, Chang-Hwan;Lee, Myoung Hee;Jung, Chan-Sik;Son, Jeong-Hee;Park, Keum-Yong
    • Korean Journal of Breeding Science
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    • v.42 no.5
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    • pp.507-514
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    • 2010
  • Peanut(Arachis hypogaea L.) is one of the major oilseed crops. The peanut oil consists of palmitic, oleic and linoleic acids, which are present at levels of 10%, 36-67% and 15-43%, respectively. High oleate mutant of peanut F435 contains 80% oleate and as little as 2% linoleate in seed oil. Previous study indicated that delta 12 fatty acid desaturase is a major enzyme controlling the oleate content in seeds of oilseed crops. F435 sequence alignment of their coding regions disclosed that an extra A(adenine) was inserted at the position +2,823 bp of delta 12 fatty acid desaturase gene. This study was to develop molecular marker (SNP marker) co-segregating with the high oleate trait. Chopyeong ${\times}$ F435 $F_2$ 41 population were investigated using molecular marker and fatty acid assay (NIR and gas chromatography). Finally, this marker segregates Chopyeong type 26 lines, heterotype 9 lines and F435 type 6 lines. These results in our study suggested that SNP marker conform fatty acid assay.

Antioxidant Activity and Whitening Efficacy of Makgeolli Fractions (막걸리 분획물의 항산화활성 및 미백효능)

  • Park, Kyung-won;Kwak, Da-hee;Kwon, Hye-Jin
    • Journal of Digital Convergence
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    • v.15 no.10
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    • pp.571-577
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    • 2017
  • This study aimed at evaluating the potential of makgeolli, which is widely consumed as beverage, as a functional cosmetic ingredient, based on analyses on its antioxidant activity and tyrosinase activity inhibitory effect. The sample was extracted by concentrating the suspension obtained after adding MeOH (3 L) to the residue of a commercial makgeolli and then adding EtOAc (3 L) to the concentrate, which was subjected to fractionation. The upper layer of the fractions was used as the final sample. In MTT assay assessments, no cytotoxicity was observed at a concentration range of 10 to $1,000{\mu}g/mL$; the antioxidant activity of the extract showed a concentration-dependent tendency and it had a high activity with an $EC_{50}$ of 7.008 mg/mL. Also, in a tyrosinase activity inhibitory effect assessment, the extract showed an $IC_{50}$ value of 39.22 mg/mL. These results confirmed that this sample has potential as a functional cosmetic ingredient.

Antioxidant Activities and Whitening Effect from Lindera obtusiloba BL. Extract (생강나무 추출물의 항산화 활성과 미백효과)

  • Bang, Chae-Young;Won, Eun-Kyung;Park, Kuen-Woo;Lee, Gwang-Won;Choung, Se-Young
    • YAKHAK HOEJI
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    • v.52 no.5
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    • pp.355-360
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    • 2008
  • In this study we investigated antioxidant activity of against several free radicals and skin whitening effect of 70% ethanol extract (leaf extracts and branch/stem mixed) of Lindera obtusiloba BL. Antioxidant activity was assessed by DPPH, superoxide radical and hydroxyl radical assays. The Lindera obtusiloba BL. extract had antioxidant activity dose dependently with an ${IC}_{50}$ value of 243.14 and 181.10 ${\mu}g$/ml for DPPH, 165.77 and >1500 ${\mu}g$/ml for non-enzymatic system of superoxide radical assay, 35.47 and >100 ${\mu}g$/ml for enzymatic system of superoxide radical assay, 1.21 mg/ml for hydroxyl radical assay. In addition we tested tyrosinase inhibition activity and melanin contents on B16 melanoma F10. B16 melanoma cell was treated by such sample as 1, 5, 10 and 50 ${\mu}g$/ml for 72 hr and tyrosinase inhibition was tested. Melanogenesis was inhibited to 22% at the dose of 50 ${\mu}g$/ml and tyrosinase was inhibited to 45.2% at the same dose. In conclusion Lindera obtusiloba BL had potent antioxidant activity and inhibitory activity of tyrosinase and melanin formation. It could be developed as the health functional food and functional cosmetic resources.

Inhibitory Effects on Oral Microbial Activity and Production of Lipopolysaccharides-Induced Pro-Inflammatory Mediators in Raw264.7 Macrophages of Ethanol Extract of Perilla flutescens (L.) Britton

  • Jeong, Moon-Jin;Lim, Do-Seon;Lee, Myoung-Hwa;Heo, Kyungwon;Kim, Han-Hong;Jeong, Soon-Jeong
    • Journal of dental hygiene science
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    • v.20 no.4
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    • pp.213-220
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    • 2020
  • Background: The leaves of Perilla frutescens, commonly called perilla and used for food in Korea, contain components with a variety of biological effects and potential therapeutic applications. The purpose of this study was to identify the components of 70% ethanol extracted Perilla frutescens (EEPF) and determine its inhibitory effects on oral microbial activity and production of nitric oxide (NO) and prostaglandin E2 (PGE2) in lipopolysaccharides (LPS)-stimulated Raw264.7 macrophages, consequently, to confirm the possibility of using EEPF as a functional component for improving the oral environment and preventing inflammation. Methods: One kg of P. frutescens leaves was extracted with 70% ethanol and dried at -70℃. EEPF was analyzed using high-performance liquid chromatography analysis, and antimicrobial activity against oral microorganisms was revealed using the disk diffusion test. Cell viability was elucidated using a methylthiazolydiphenyl-tetrazolium bromide assay, and the effect of EEPF on LPS-induced morphological variation was confirmed through microscopic observation. The effect of EEPF on LPS-induced production of pro-inflammatory mediators, NO and PGE2 was confirmed by the NO assay and PGE2 enzyme-linked immunosorbent assay. Results: The main component of EEPF was rosemarinic acid, and EEPF showed weak anti-bacterial and anti-fungal effects against microorganisms living in the oral cavity. EEPF did not show toxicity to Raw264.7 macrophages and had inhibitory effects on the morphological variations and production of pro-inflammatory mediators, NO and PGE2 in LPS-stimulated Raw264.7 macrophages. Conclusion: EEPF can be used as a functional material for improving the oral environment through the control of oral microorganisms and for modulating inflammation by inhibiting the production of inflammatory mediators.

The anti-tumor efficacy of 20(S)-protopanaxadiol, an active metabolite of ginseng, according to fasting on hepatocellular carcinoma

  • Li, Wenzhen;Wang, Yifan;Zhou, Xinbo;Pan, Xiaohong;Lu, Junhong;Sun, Hongliu;Xie, Zeping;Chen, Shayan;Gao, Xue
    • Journal of Ginseng Research
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    • v.46 no.1
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    • pp.167-174
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    • 2022
  • Background: 20(S)-protopanaxadiol (20(S)-PPD), one of the main active metabolites of ginseng, performs a broad spectrum of anti-tumor effects. Our aims are to search out new strategies to enhance anti-tumor effects of natural products, including 20(S)-PPD. In recent years, fasting has been shown to be multi-functional on tumor progression. Here, the effects of fasting combined with 20(S)-PPD on hepatocellular carcinoma growth, apoptosis, migration, invasion and cell cycle were explored. Methods: CCK-8 assay, trypan blue dye exclusion test, imagings photographed by HoloMonitorTM M4, transwell assay and flow cytometry assay were performed for functional analyses on cell proliferation, morphology, migration, invasion, apoptosis, necrosis and cell cycle. The expressions of genes on protein levels were tested by western blot. Tumor-bearing mice were used to evaluate the effects of intermittent fasting combined with 20(S)-PPD. Results: We firstly confirmed that fasting-mimicking increased the anti-proliferation effect of 20(S)-PPD in human HepG2 cells in vitro. In fasting-mimicking culturing medium, the apoptosis and necrosis induced by 20(S)-PPD increased and more cells were arrested at G0-G1 phase. Meanwhile, invasion and migration of cells were decreased by down-regulating the expressions of matrix metalloproteinase (MMP)-2 and MMP-9 in fasting-mimicking medium. Furthermore, the in vivo study confirmed that intermittent fasting enhanced the tumor growth inhibition of 20(S)-PPD in H22 tumor-bearing mice without obvious side effects. Conclusion: Fasting significantly sensitized HCC cells to 20(S)-PPD in vivo and in vitro. These data indicated that dietary restriction can be one of the potential strategies of chinese medicine or its active metabolites against hepatocellular carcinoma.

Anti-Oxidant and Anti-Inflammatory Complex Health Functional Analysis of Chinese Resource Plant Boehmeria, Carlemannia and Clausena (중국 자원식물 보메리아, 칼라마니아, 클로세나의 항노화 항염 복합 건강관리 기능성 연구)

  • Jin Mun;Jung Min Lee;Je Heon Lee;Dong Hyun Lee;Seo Yeon Choi;Ye Lim Jung;Kyung-A Choi
    • Proceedings of the Plant Resources Society of Korea Conference
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    • 2023.04a
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    • pp.52-52
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    • 2023
  • Ageing has been known to be deeply related to oxidative activities. Furthermore, inflammation is a response initiated by conditions such as infection and injury. It has been known that complex connections occur between oxidative stress and the inflammatory response, however underlying mechanism needs to be investigated. Chinese resource plants have been widely used as functional food and medicine for a long time, however it is not studied well how many of these resource plants work. We first decided testing anti-oxidant and anti-inflammatory activities with 95% ethanol extracts of the chinese resource plants, Boehmeria pilosinscula (Blume) Hassk., Carlemannia tetragona Hook f. and Clausena emarginata C.C.Huang. To measure anti-oxidant activity, we performed DPPH assay in Raw264.7 cells with 95% ethanol extracts. Clausena extracts showed significantly higher anti-oxidant activity than those of Boehmeria and Carlemannia. We then performed Nitric Oxide assay to measure the inflammation suppression levels with 95% ethanol extracts of these plants. Clausena extracts showed significantly higher suppression of nitric oxide production than those of Boehmeria and Carlemannia. This indicates that inflammation levels are significantly reduced by Clausena. After measuring anti-oxidant and anti-inflammatory activities, we then performed MTT assay to measure the cell survival rate of Raw264.7 cells treated with these extracts. Boehmeria showed much more cell survival rate than Carlemannia and Clausena. Taken together, this result suggests that Clausena extracts have more anti-oxidant and antiinflammatory activities than Boehmeria and Carlemannia, while Boehmeria extracts have more cell survival rate than Carlemannia and Clausena.

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The effects of ATO-ALL on skin regeneration in human epidermal keratinocytes, HaCaT cells (ATO-ALL이 인간유래 피부 각질형성세포인 HaCaT 세포에서 피부재생에 미치는 효과)

  • Shin, Younmin;Lee, Won-Kil;Kim, Seung Hyung;Choi, Jeong June
    • Journal of Haehwa Medicine
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    • v.28 no.2
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    • pp.41-47
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    • 2019
  • Objectives : Atopic dermatitis is an irritable skin disease accompanying rash and itching leading to impaired skin barrier. ATO-ALL is an ethanol extract of natural products comprising 12 herbs and effective on atopic dermatitis. In this study, we aimed to propose that the effect of ATO-ALL on skin regeneration in human keratinocyte cell line, HaCaT cells. Methods : To evaluate the skin regenerating effects of ATO-ALL, scratch wound healing assay, bromodeoxyuridine (BrdU) assay, and propidum iodide (PI) assay were performed using cultured HaCaT cell line. Result : Scratch wound healing assay showed that ATO-ALL was able to enhance the gap filling activity more than 2-fold at 7 ppm concentration compared with control group. BrdU assay demonstrated that ATO-ALL treatment increased the de novo cell proliferation in a dose-dependent manner. Finally, PI assay indicated that the cell cycle of HaCaT cells was modulated by ATO-ALL treatment. Conclusions : These results suggested that ATO-ALL may have skin regenerating effects by increasing cell proliferation via cell cycle regulation. Taken together, ATO-ALL is supposed to have a potential on regeneration of damaged skin or functional disease including atopic dermatitis.

Properties of Fucoidan as Raw Materials of Water-holding Cream and Cosmetics (후코이단의 보습제 및 화장품재료로서 특성)

  • Cha, Sung Hwan;Lee, Jung Shik;Kim, Young Suk;Kim, Donguk;Moon, Jae Cheon;Park, Kwonpil
    • Korean Chemical Engineering Research
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    • v.48 no.1
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    • pp.27-32
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    • 2010
  • Properties of fucoidan used for functional cosmetics agents were studied. Fucoidan was extracted from Undaria pinnatifida sporophylls. To test possibility of fucoidan as a cosmetics material, water-holding property measurement, DPPH free radical scavenging assay and MTT assay were done. Water-holding property of fucoidan was higher than that of hyaruronic acid, which is known as the one of the best water-holding material. The water-holding strength of fucoidan slightly increase as molecular weight of fucoidan decrease. Fucoidan showed high stability from MTT assay and good anti-oxidation property from DPPH assay. To evaluate the effect of water-holding property and anti-alergy property of fucoidan on the atopic dermatitis(AD), 46 AD patients were treated with fucoidan cream. After 6 weeks treatment, Investigation Global Assessment(IGA) scores decreased from 3.04 to 2.15, that is fucoidan cream had a 39.8% benefit effect on atopic dermatitis.