• Korean Journal of Food Science and Technology
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• v.53 no.2
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• pp.204-212
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• 2021

Resazurin, an oxidized blue dye, is reduced to resorufin, showing a peak absorbance change and emitting fluorescence due to the metabolic activity of living cells. In this study, the growth of Lactobacillus rhamnosus GG (LGG) and the redox potential of living probiotic bacteria were evaluated based on changes in the absorbance or fluorescence of resazurin. Fluorescence analysis is a more accurate and sensitive method for quantifying viable LGG than the colorimetric absorbance measurement of resazurin change. Fluorescence measurement could detect LGG of 6.5-9.5 log CFU/mL within 30 min with R2=0.99. No significant effect of further reduction of resorufin to dihydroresorufin by LGG was observed. Various probiotics showed different resazurin-reducing activities, and L. kimchicus had the highest reducing activity among the six probiotics tested. These findings suggests that fluorescence measurement in a resazurin-based assay is useful for analyzing bacterial growth and the redox potential of living probiotics.

• Proceedings of the Plant Resources Society of Korea Conference
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• 2019.10a
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• pp.64-64
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• 2019

Screening and identification of genetic resources based on their phytoconstituents and morphological characters potentially provide baseline data for researchers, breeders, and nutraceutical companies who wish to formulate a nutrient-dense diet and health beneficial supplement. Thus, we evaluated the amount of total phenolic content and major phenolic compounds; examined if phenolic compounds could be used as distinguishing factors for perilla genetic resources; and investigated the relation between some quantitative and qualitative morphological characters with the contents of phenolic compounds in 360 accessions obtained from National Agrobiodiversity Center gene bank, Jeonju, Korea. Total phenolic content (TPC) was estimated using Folin-Ciocalteu colorimetric assay. Individual phenolic compounds were determined using an Ultra-High Performance Liquid Chromatography system equipped with Photodiode Array detector. Considerable variations were observed in TPC (7.99 to 117.47 mg GAE/g DE), rosmarinic acid (RA) (ND to 19.19 mg/g DE), caffeic acid (CA) (ND to 0.72 mg/g DE), apigenin-7-O-diglucuronide (ADG) (ND to 1.24 mg luteolin equivalent (LUE)/g DE), scutellarein-7-O-glucuronide (SG) (ND to 4.32 mg LUE/g DE), and apigenin-7-O-glucuronide (AG) (ND to 1.60 mg LUE/g DE). RA was the most dominant phenolic compound in most accessions (95.3%) followed by SG. The adaxial leaf color was light green, green and dark green in 13.8%, 65.0%, and 21.1 % of the accessions, respectively. 78.8% of the accessions had light green color at the abaxial side with the remaining being described as green. Most of the accessions (96.9%) were cordate shape, the remaining being eclipse. Intensities of green pigment at abaxial and adaxial leaf surfaces were correlated with contents of individual phenolic compounds and TPC whereas leaf length and width had no correlation with TPC, CA and RA, and negatively correlated with ADG, AG, and SG. Leaf shape was not related with content of phenolic compounds, color of leaves, or the length or width of leaves. Accessions IT57426, IT157434, IT267710, and IT267712 which contained relatively high contents of TPC and major phenolic compounds (RA and SG) could be used for further research in breeding and bioassay test. Our study result showed the contents of total phenolics and individual phenolic compounds along with the morphological characters could be useful distinguishing factors for perilla genetic resources.

• The Korea Journal of Herbology
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• v.33 no.3
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• pp.37-43
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• 2018

Objectives : Butylated hydroxytoluene (BHT) and butylated hydroxyanisole (BHA) were well known as anti-oxidant, but they were limited to use because of toxicity. So, many studies are being done to develope natural anti-oxidant. Total phenolic and flavonoid contents along with total antioxidant capacity of the ethanolic extract of Plantaginis Herba (PH) were evaluated to explore the reliable and potential sources of novel natural antioxidants. Methods : Total polyphenol contents and total flavonoid contents in PH ethanol extract were determined by colorimetric method. And DPPH(1.1-diphenyl-2-picrylhydrazyl), ABTS(2'-azino-bis (3-ethylbenzothiazoline-6-Surfonicacid)) free radical scavenging capacity and reducing power inhibition activities of PH ethanol extract were measured at 100, 500, 1000, $5000{\mu}g/m{\ell}$ concentrations by spectrometric assay. Results : The total polyphenol contents and total flavonoid contents of the extract were 161.99 mg/g, 144.05 mg/g, respectively. Also, DPPH, ABTS free radical scavenging capacity and reducing power of PH ethanol extract in treated concentrations (100, 500, 1000, $5000{\mu}g/m{\ell}$) increased dose dependently. In particular, DPPH free radical scavenging capacity of PH ethanol extract from $500{\mu}g/m{\ell}$ was significantly increased compared to positive control (BHA). ABTS free radical scavenging capacity of PH ethanol extract from $1000{\mu}g/m{\ell}$ was significantly higher than BHA. Also, reducing power showed that PH ethanol extract from $500{\mu}g/m{\ell}$ was significantly increased compared to BHA. Conclusions : These results suggest that PH ethanol extract has effects to scavenge free radicals, thus PH has potential and applicable benefits for development of materials and products to have anti-oxidation functions.

• Journal of Nutrition and Health
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• v.55 no.1
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• pp.36-46
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• 2022

Purpose: Quercetin is a polyphenolic flavonoid abundant in many fruits and vegetables. It has potential health-beneficial properties, such as antioxidant, anti-obesity, anti-cancer, anti-diabetic and anti-inflammatory effects. The purpose of this study was to investigate whether the lipid metabolism improvement effect of quercetin affected the regulation of AMP-activated protein kinase (AMPK) activity and microRNA (miR)-21 expression in the liver of mice fed a high-cholesterol diet. Methods: Male C57BL/6J mice were fed with normal diet, quercetin-free diet and diets containing 0.05% or 0.1% quercetin for six weeks. Hypercholesterolemia was induced by adding 1% cholesterol and 0.5% cholic acid to all diets. Serum and liver triglyceride (TG), and total cholesterol (TC) concentrations were analyzed using a commercial enzymatic colorimetric kit. AMPK activity was quantified using an AMPK kinase assay kit. The levels of miR-21 and genes involved in lipid metabolism were measured by real-time quantitative polymerase chain reaction. Results: Supplementation of quercetin reduced serum and hepatic TG and TC levels without changing body weight and food intake. Dietary quercetin significantly inhibited the mRNA levels of hepatic sterol-regulatory element binding protein-1c, acetyl-CoA carboxylase 1 and fatty acid synthesis, which are involved in hepatic lipogenesis. Dietary quercetin enhanced AMPK activity and suppressed miR-21 expression, promoting hepatic lipid accumulation. Conclusion: These results suggest that the lipid-lowering effect of quercetin on the serum and liver of mice may be partially mediated by the regulation of lipogenic gene expression, AMPK activity and miR-21 expression in the liver of mice fed a high-cholesterol diet.

• Journal of the Korean Society of Food Science and Nutrition
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• v.42 no.2
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• pp.161-167
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• 2013

The aim of this study was to investigate the protective effects of methanolic extract from perilla (Perilla frutescens Britt var. japonica) leaves (PLME) on oxidative injury from hydrogen peroxide ($H_2O_2$) in human HaCaT keratinoctyes. Cells were co-incubated with various concentrations (0~200 ${\mu}g/mL$) of PLME for 24 hr, and then exposed to $H_2O_2$ (500 ${\mu}M$) for 4 hr. $H_2O_2$ significantly decreased cell viability (p<0.05). However, PLME provided protection from $H_2O_2$-induced HaCaT cell oxidation in a dose-dependent manner. To further investigate the protective effects of PLME on $H_2O_2$-induced oxidative stress in HaCaT cells, the cellular levels of lipid peroxidation, and antioxidant enzymes (including superoxide dismutase (SOD), glutathione peroxidase (GSH-px) and catalase (CAT)) were measured. PLME decreased cellular levels of lipid peroxidation, and also increased the activities of antioxidant enzymes. In addition, the antioxidant activities of PLME were also determined by DPPH and hydroxyl (${\cdot}OH$) radical scavenging assay, and major antioxidant compounds of PLME were measured by colorimetric methods. DPPH and ${\cdot}OH$ radical scavenging activities of PLME increased in a dose dependent manner and was similar to the DPPH scavenging activity of ascorbic acid at 50 ${\mu}g/mL$; however PLME activities were stronger than ascorbic acid (50 ${\mu}g/mL$) in the ${\cdot}OH$ scavenging assay. The amounts of antioxidant compounds, including total polyphenolics, total flavonoids, and total ascorbic acid from PLME were $52.2{\pm}1.1$ mg gallic acid (GAE)/g, $33.7{\pm}4.7$ mg rutin (RUE)/g, and $17.0{\pm}0.5$ mg ascorbic acid (AA)/g, respectively. These results suggest that PLME has a strong free radical-scavenging activity and a protective effect against $H_2O_2$-induced oxidative stress in the keratinocytes.

• Journal of Korean Ophthalmic Optics Society
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• v.20 no.2
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• pp.237-246
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• 2015

Purpose: The present study was conducted to reveal the correlation of structural denaturation and decrease of enzyme activity when the antioxidative enzymes, superoxide dismutase (SOD) and catalase (CAT) were repeatedly exposed to UV-B, and further investigate whether the denaturation and inactivation of those enzymes can be effectively blocked by using UV-inhibitory RGP lens. Methods: Each enzyme solution was prepared from the standardized SOD and CAT, and repeatedly exposed to UV-B of 312 nm for 30 minutes, 1 hour and 2 hours a day over 1, 2, 3, 4 and 5 days. Structural denaturation of SOD and CAT induced by repeat UV-B irradiation was confirmed by the electrophoretic analysis, and their enzyme activity was determined by the colorimetric assay using the proper assay kit. At that time, the change in structure and activity of the antioxidant enzymes directly exposed to UV-B was compared to the case that UV-B was blocked by UV-inhibitory RGP lens. Results: SOD exposed repeatedly to UV-B showed the polymerization pattern in the electrophoretic analysis when it repeatedly exposed for 30 min a day, however, the change of its activity was less than 10%. On the other hand, CAT repeatedly exposed to UV-B reduced size and density of the electrophoretic band which indicated a structure denaturation, and its activity was significantly decreased. In the case that the repeat exposure time was longer, CAT activity was completely lost even though some enzyme band occurred in the electrphoretic analysis. In addition, the degeneration of CAT due to UV-B irradiation was inhibited to some extent by using RGP lens with a UV-B blocking of 63.7%, however, it was not completely inhibited. Conclusions: From these results, it was revealed that the structural denaturation of antioxidative enzymes was not perfectly correlated with the reduction in enzyme activity according to the type of enzyme. It is recommended to minimize the exposure time to UV when wearing contact lens, or wear the contact lenses having UV blocking rate of the FDA Class I blocker or the sunglasses having equivalent UV-blocking rate for reducing the damage of antioxidative enzymes induced by UV.

• Journal of Nutrition and Health
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• v.54 no.6
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• pp.618-630
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• 2021

Purpose: The ginger rhizome (Zingiber officinale) is widely cultivated as a spice for its aromatic and pungent components. One of its constituents, 6-hydroxydopamine (6-OHDA) is usually thought to cross the cell membrane through dopamine uptake transporters, and induce inhibition of mitochondrial respiration and the generation of intracellular reactive oxygen species (ROS). This study examines the neuroprotective effect and acetylcholinesterase (AChE) inhibitory activity of fermented ginger extracts (FGEs) on 6-OHDA induced toxicity in SH-SY5Y human neuroblastoma cells. Methods: Ginger was fermented using 2 species of Bacillus subtilis, with or without enzyme pretreatment. Each sample was extracted with 70% ethanol. Neurotoxicity was assessed by applying the EZ-Cytox cell viability assay and by measuring lactic dehydrogenase (LDH) release. Morphological changes of apoptotic cell nuclei were observed by Hoechst staining. Cell growth and apoptosis of SH-SY5Y cells were determined by Western blotting and enzyme activity analysis of caspase-3, and AChE enzymatic activity was determined by the colorimetric assay. Results: In terms of cell viability and LDH release, exposure to FGE showed neuroprotective activities against 6-OHDA stimulated stress in SH-SY5Y cells. Furthermore, FGE reduced the 6-OHDA-induced apoptosis, as determined by Hoechst staining. The occurrence of apoptosis in 6-OHDA treated cells was confirmed by determining the caspase-3 activity. Exposure to 6-OHDA resulted in increased caspase-3 activity of SH-SY5Y cells, as compared to the unexposed group. However, pre-treatment with FGE inhibited the activity of caspase-3. The neuroprotective effects of FGE were also found to be caspase-dependent, based on reduction of caspase-3 activity. Exposure to FGE also inhibited the activity of AChE induced by 6-OHDA, in a dose-dependent manner. Conclusion: Taken together, our results show that FGE exhibits a neuroprotective effect in 6-OHDA treated SH-SY5Y cells, thereby making it a potential novel agent for the prevention or treatment of neurodegenerative disease.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.18
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• pp.7849-7855
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• 2014

Purpose: To investigate the effect of deacetylase inhibitory trichostatin A (TSA) on anti HepG2 liver carcinoma cells and explore the underlying mechanisms. Materials and Methods: HepG2 cells exposed to different concentrations of TSA for 24, 48, or 72h were examined for cell growth inhibition using CCK8, changes in cell cycle distribution with flow cytometry, cell apoptosis with annexin V-FTIC/PI double staining, and cell morphology changes under an inverted microscope. Expression of ${\beta}$-catenin, HDAC1, HDAC3, H3K9, CyclinD1 and Bax proteins was tested by Western blotting. Gene expression for ${\beta}$-catenin, HDAC1and HDAC3 was tested by q-PCR. ${\beta}$-catenin and H3K9 proteins were also tested by immunofluorescence. Activity of Renilla luciferase (pTCF/LEF-luc) was assessed using the Luciferase Reporter Assay system reagent. The activity of total HDACs was detected with a HDACs colorimetric kit. Results: Exposure to TSA caused significant dose-and time-dependent inhibition of HepG2 cell proliferation (p<0.05) and resulted in increased cell percentages in G0/G1 and G2/M phases and decrease in the S phase. The apoptotic index in the control group was $6.22{\pm}0.25%$, which increased to $7.17{\pm}0.20%$ and $18.1{\pm}0.42%$ in the treatment group. Exposure to 250 and 500nmol/L TSA also caused cell morphology changes with numerous floating cells. Expression of ${\beta}$-catenin, H3K9and Bax proteins was significantly increased, expression levels of CyclinD1, HDAC1, HDAC3 were decreased. Expression of ${\beta}$-catenin at the genetic level was significantly increased, with no significant difference in HDAC1and HDAC3 genes. In the cytoplasm, expression of ${\beta}$-catenin fluorescence protein was not obvious changed and in the nucleus, small amounts of green fluorescence were observed. H3K9 fluorescence protein were increased. Expression levels of the transcription factor TCF werealso increased in HepG2 cells following induction by TSA, whikle the activity of total HDACs was decreased. Conclusions: TSA inhibits HDAC activity, promotes histone acetylation, and activates Wnt/${\beta}$-catenin signaling to inhibit proliferation of HepG2 cell, arrest cell cycling and induce apoptosis.

• Journal of Periodontal and Implant Science
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• v.31 no.4
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• pp.833-846
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• 2001

Recently, use of natural medicine is getting more attention, and some of them are believed to be effective in　the treatment of periodontitis. Among them, the seeds of safflower(Carthamus tinctrorius L.) have been proven to be effective through its use in bone diseases such as fracture and osteoporosis. During the last few years, studies using the seeds of safflower gown in Korea have been active, and it has been reported that safflower seed extract increase the proliferation and the alkaline phosphatase(ALP) activity of human periodontal ligament fibroblast(hPDLF), osteoblast, and that they promote the mineralization process. In animal studies, when safflower seed extract were administered orally new bone formation was promoted. Recently, in an effort to find out the most effective osteogenic components, among many components of the safflower seed, various safflower seed fraction extracts were obtained by multistep extraction of the safflower components using various solvents. Among these, saf-M-W fraction extracted by methanol and water was most effective in increasing osteogenic potential of osteoblasts. In this study, the effect of safflower seed fraction extract, saf-M-W, on the growth and differentiation of hPDLF and MC3T3-E1 cell was investigated. The toxicity of saf-M-W on both cells was measured using M'IT(3-(4,5dimethylthiazol-2-y1)-2,5-diphenyl tetrazolium bromide) test, and ALP activity was measured using the colorimetric assay of hPDLF. In addition, in MC3T3-El cells, the expression of ALP, bone sialoprotein(BSP) mRNA was observed using Northern blot, and the mineralized nodule formation Was observed using von Kossa stain and phase-contrast microscope. 1. In concentrations below $10{\mu}g/ml$, saf-M-W didn't show any toxicity on hPDLF and MC3T3-El cell. 2. The change in saf-M-W concentration had no effect on the ALP activity of hPDLF. 3. In MC3T-E1 cells, mRNA expressions of ALP and BSP were greater in the experimental group treated with $10{\mu}g/ml$ concentration of saf-M-W compared with the control group. 4. In MC3T3-El cells, abundance of mineralized nodules were formed in the experimental group treated with $10{\mu}g/ml$ Concentration of saf-M-W, while no mineralized nodule was formed in the control group. These results suggest that safflower seed fraction extract, saf-M-W. didn't show any toxicity on hPDLF and MC3T3-E1 cell at concentrations below $10{\mu}g/ml$ and effectively enhanced the differentiation and osteogenic potential of MC3T3-El cell.

• The Korean Journal of Nuclear Medicine
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• v.29 no.3
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• pp.332-342
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• 1995

This study was designed to evaluate the effects of various factors on the therapeutic effect of the I-131 labeled anti-carcinoembryonic antigen monoclonal antibody(anti-CEA antibody). Tetrazolium-based colorimetric assay (MTT) was used to compare in vitro cytotoxicity of 3 Korean colon cancer cell lines (SNU-C2A, SNU-C4, SNU-C5) for selection of proper 2 cell lines in this study. The changes of the size of tumor which was xenografted to nude mice (balb/c nu/nu) were compared in 4 groups (group treated I-131 labeled anti-CEA antibody, group treated with non-radiolabeled anti-CEA antibody, group treated with I-131 labeled anti-human chorionic gonadotropin monoclonal antibody (anti-hCG antibody) as nonspecific antibody, and group injected with normal saline as a control). Immunohistochemical staining and in vivo autoradiography were performed after excision of the xenografted tumor. The results were as below mentioned. The in vitro cytotoxic effect of I-131 labeled anti-CEA antibody is most prominent in SNU-C5 cell line between 3 cancer cell lines. The changes of xenografted tumor size in both SNU-C4 and SNU-5S cell tumors at the thirteenth day after injection of the antibodies were smallest in the group treated with I-131 labeled anti-CEA antibody (SNU-C4/SNU-C5; 324/342%) comparing with other groups, group treated with anti-CEA antibody (622/660%), group treated with I-131 anti-hCG antibody (538/546%), and control group(1030/724%)(P<0.02 in SNU-C4 and P<0.1 in SNU-C5 at the 13th day after injection of antibodies). On the thirteenth day after injection of the antibodies nude mice were sacreficed to count the radiouptake of tumor and to check the changes of tumor size. Correlations between radiouptake and change of tumor size were calculated in each groups and significant negative correlation was only obtained in the group treated with I-131 anti-CEA antibody (p<0.05). There were no correlations between antigenic expression of carcinoembryonic antigen and distribution of anti-CEA antibody in both SNU-C4 and SNU-C5 cell tumors on immunoperoxidase staining. On in vivo autoradiography the distributions of anti-CEA antibody were heterogeneous and the intensities of binding were various in SNU-C4 and SNU-C5 cell tumors. It is concluded that I-131 labeled tumor-specific monoclonal antibody, anti-CEA antibody is effective in suppressing the xenografted tumor growth and the effect is influenced by sensitivity of tumor cell itself to the radiolabeled antibody and other local factors instead of specificity of antibody.