Inhibitory Effects of Flavonoids Isolated from the Leaves of Stewartia koreana on Nitric-oxide Production in LPS-stimulated RAW 264.7 Cells

Seung-Su Lee(이승수),Myun-Ho Bang(방면호),Se-Ho Park(박세호),Dae-kyun Chung(정대균),Seun-Ah Yang(양선아) 한국생명과학회 2018 생명과학회지 Vol.28 No.5

노각나무(Stewartia koreana) 잎 에틸아세테이트 분획으로부터 quercetin (1), quercitrin (2), hyperin (3), quercetin-3-O-(6""-O-galloyl)-β-D-galactopyranoside (4), kaempferol-3-o-[2"",6""-di-o-(trans-p-coumaroyl)]-β-D-glucopyranoside (5)의 5종의 플라보노이드를 분리하였으며, 이들 5종 성분의 염증 반응에 대한 활성을 분석하기 위하여 LPS를 처리한 대식세포에서 산화질소(NO) 생성 억제활성을 측정하였다. 이들 5종 성분 중 compound 4, 5는 노각나무에서 처음으로 분리된 것으로 항염증 활성에 대한 보고도 없다. 분광분석법으로 확인된 노각나무 잎 유래성분들은 LPS 처리한 대식세포의 NO 생성을 유의적으로 저해하였으며, 특히 kaempferol-3-o-[2"",6""-di-o-(transp-coumaroyl)]-β-D-glucopyranoside (5)는 가장 강한 억제효과(17.17%, 5.0%, 3.92%, 6.32% and 63.35% inhibition of compound 1, 2, 3, 4 and 5 at 10 μg/ml)를 나타냈다. 또한, 이러한 NO 생성 억제효과는 inducible nitric oxide synthase(iNOS) 단백질 발현 억제를 통한 것으로 나타났다. 따라서, 본 연구에서 새로 분리된 플라보놀인 kaempferol-3-o-[2"",6""-di-o-(trans-p-coumaroyl)]-β-D-glucopyranoside (5)는 노각나무 잎의 항염증 활성을 나타내는 주요 물질로 사료된다. Five phenolic compounds were isolated from the ethyl acetate fraction of leaves from Stewartia koreana, and their nitric-oxide (NO) inhibitory activities were measured to identify the major active constituents responsible for the efficacy of the extract against inflammatory reactions. These five compounds were quercetin (1), quercitrin (2), hyperin (3), quercetin-3-O-(6""-O-galloyl)-β-D-galactopyranoside (4), and kaempferol 3-O-[2"",6""-di-O-(trans-p-coumaroyl)]-β-D-glucopyranoside (5). Among the separated compounds in the EtOAc fraction, compounds 4 and 5 were isolated for the first time, and no study has yet reported their anti-inflammatory effects. The compounds were identified by spectroscopic analysis, and the isolated compounds showed significant NO inhibitory effects in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. Compound 5 showed the most potent inhibitory effect (63.35% inhibition) against LPS-induced NO production compared to that of compound 1 (17.17%), compound 2 (5.0%), compound 3 (3.92%), and compound 4 (6.32%) at 10 μg/ml concentration. NO production was inhibited by suppressing the protein expression of inducible nitric-oxide synthase in LPS-stimulated RAW 264.7 macrophages. These results indicate that kaempferol 3-O-[2"",6""-di-O-(trans-p-coumaroyl)]-β-D-glucopyranoside might be the major active compound responsible for the anti-inflammatory effects of S. koreana.

노각나무(Stewartia koreana, 차나무과)의 분류학적 재검토

권혜진,송호경,김무열 한국식물분류학회 2008 식물 분류학회지 Vol.38 No.3

Morphological and molecular data from the nuclear ribosomal ITS regions were examined to reevaluate the taxonomic position of the Korean endemic species, Stewartia koreana Nakai ex Rehder (Theaceae). Comparing to its morphologically closely related S. pseudocamellia Maxim, S. koreana has diagnostic characters including a larger leaf blade (10 cm length and 5.5 cm wide) and a longer pedicel (2.5 cm length). In addition, S. koreana formed a different clade from S. pseudocamellia in the ITS trees. The morphological and ITS sequence data supported S. koreana as a Korean endemic species distinct from the closely related species, S. pseudocamellia. 한국 특산종인 노각나무(Stewartia koreana Nakai ex Rehder)의 분류학적 위치를 재검토하기 위해, 노각나무와 근연 분류군의 외부형태와 ITS 염기서열을 조사하였다. 노각나무는 일본에 분포하는 S. pseudocamellia와 외부 형태적으로 매우 유사하였으나, 노각나무는 잎이 대형이고 소화경의 길이가 2.5 cm 이상인 점에서 S. pseudocamellia와 구별되었다. ITS 염기서열을 분석한 결과 노각나무는 S. pseudocamellia와 별개의 분계조를 형성하였다. 따라서 외부형태 및 ITS 염기서열 결과는 노각나무가 S. pseudocamellia와 다른 별개의 한국 특산종임을 지지해 주었다.

Stewartia koreana extract stimulates proliferation and migration of human endothelial cells and induces neovasculization in vivo

Lee, Tae Hoon,Lee, Guy Wilhem,Kim, Chan Woo,Bang, Myun-Ho,Baek, Nam-In,Kim, Sung-Hoon,Chung, Dae Kyun,Kim, Jiyoung John Wiley Sons, Ltd. 2010 Phytotherapy research Vol.24 No.1

<P>Angiogenesis, the growth of new blood vessels from preexisting vasculature, plays an important role in physiological and pathological processes such as embryonic development and wound healing. This study investigated the effects of methanol extracts of Stewartia koreana leaves (SKE) on angiogenesis. Stewartia koreana significantly promoted the proliferation and migration of human umbilical vein endothelial cells in a dose-dependent manner. The SKE induced endothelial cell proliferation in the range of 50 µg/mL without cytotoxicity. Treatment of HUVECs resulted in the activation of the mitogen-activated protein kinases that was correlated with endothelial cell proliferation and migration. SKE also stimulated angiogenesis in a chick chorioallantoic membrane assay, demonstrating promotion of new blood vessel formation in vivo. Local administration of SKE onto skin punched wounds resulted in increased von Willebrand Factor antigen, indicating that it stimulated neovasculization in the wound region. The results suggest that Stewartia koreana extracts may potentially be useful for the development of agents to accelerate vascular wound healing or to promote the growth of collateral blood vessels in ischemic tissues. Copyright © 2009 John Wiley & Sons, Ltd.</P>

노각나무 추출물이 Pseudomonas aeruginosa의 바이오필름 형성에 미치는 영향

이상균,김혜수,조수정 한국생명과학회 2023 생명과학회지 Vol.33 No.11

본 연구에서는 천연물 유래 항균소재로써 노각나무의 이용 가능성을 알아보기 위해 노각나무 가지, 줄기, 잎을 에탄올에 추출한 다음 P. aeruginosa에 대한 추출물의 항균활성을 조사하였다. 노각나무 부위별 추출물(1 mg/disc) 중 잎과 줄기 추출물이 P. aeruginosa에 대해 항균활성을 나타내었으며, 잎 추출물의 항균활성이 가장 우수하였다. P. aeruginosa에 대한 노각나무 잎 추출물의 MIC는 0.8 mg/ml였고 정균작용을 나타내었다. 노각나무 잎 추출물이 0.2-2.0 mg/ml 농도로 처리된 배양액에서 P. aeruginosa의 바이오필름 형성은 추출물의 농도가 증가할수록 농도의존적으로 억제되는 경향을 보였다. 또한 주사전자현미경으로 노각나무 잎 추출물(0.8 mg/ml)이 P. aeruginosa의 바이오필름 형성에 미치는 영향을 관찰한 결과에 의하면 추출물 대신 DMSO를 처리한 대조구에서는 세포 주변에서 EPS와 바이오필름이 관찰되었지만 추출물을 처리한 처리구의 세포 주변에서는 EPS와 바이오필름이 관찰되지 않았다. qRT-PCR을 이용하여 P. aeruginosa의 QS 관련 유전자인 lasI와 rh1I mRNA 발현 변화를 qRT-PCR로 확인한 결과, 노각나무 잎 추출물이 0.2-2.0 mg/ml의 농도로 처리된 배양액에서 lasI와 rh1I mRNA 발현은 추출물의 농도가 높아질수록 농도의존적으로 억제되는 것을 확인할 수 있었다. 이상의 결과를 종합하면 노각나무 잎 추출물은 P. aeruginosa에 대한 항균활성과 바이오필름 형성 억제능이 우수하기 때문에 천연물 유래 항균소재로써 이용 가능성이 높을 것으로 판단된다. This study was conducted to investigate the potential of Stewartia koreana as anti-microbial materials. The branches, stems and leaves of S. koreana were extracted into 70% ethanol and their antibacterial activity against P. aeruginosa was confirmed. The leaf, branch and stems extracts (1 mg/disc) showed the antibacterial activity against P. aeruginosa and leaf extracts showed higher antibacterial activities than those from branch extracts. The MIC against P. aeruginosa was 0.8 mg/ml and showed bacteriostatic action. The inhibitory effects of extract on biofilm formation and gene expression related to biofilm formation of P. aeruginosa was determined by biofilm biomass staining, SEM and qRT-PCR analysis. The biofilm biomass and cell growth of P. aeruginosa in the cultures treated with 0.2~2.0 mg/ml of S. koreana leaf extracts were significantly decreased in a concentration-dependent manner. We observed that the extract had an inhibitory effect on the formation of P. aeruginosa biofilms at concentrations of 0.8 mg/ml by SEM. qRT-PCR analysis showed that the lasI and rh1I gene expression associated to quorum sensing (QS) in the cultures treated with 0.2~2.0 mg/ml of S. koreana leaf extracts were suppressed in a concentration-dependent manner. Based on the above results, it can be concluded that S. koreana leaf extracts can be used as anti-microbial material derived from natural materials, as demonstrated by the antibacterial action and inhibition of biofilm formation of P. aeruginosa by QS inhibition.

Quantitative analysis of hyperoside and isoquercitrin in methanolic extract of Stewartia koreana leaves using HPLC-DAD

Ju-Yeong Kang,Yu Hwa Kim,Youngdae Yoon,Bong-Gyu Kim 한국응용생명화학회 2023 Journal of Applied Biological Chemistry (J. Appl. Vol.66 No.-

Since Stewartia koreana leaves are registered with the Food and Drug Administration as edible herbal materials, they are used in the development of functional foods, cosmetics, and medicines. In this study, we established an analysis method that can simultaneously analyze two indicators, hyperoside (quercetin 3-O-galactoside) and isoquercitrin (quercetin 3-O-glucoside) contained in the leaves of S. koreana using HPLC-DAD. In accordance with the Ministry of Food and Drug Safety's health functional food guidelines, the analysis method was verified for specificity, accuracy, precision, limit of quantification, and linearity. The analysis method established in this study showed more than 0.9989 of the correlation coefficient values (r2) for the calibration. The total recovery rates of isoquercitrin and hyperoside were 100.55 and 98.87% with 0.14-0.78 and 0.47-0.67% of the relative standard deviation, respectively. Therefore, it was suggested that the new analytical method would be applied to standardize raw materials and high value-added products originated from the leaves of the S. koreana in the future.

노각나무 잎과 가지 추출물의 항산화 효과

김혜수(Hye Soo Kim),박민정(Min Jeong Park),김수정(Soo Jeong Kim),김부경(Bu Kyung Kim),박준호(JunHo Park),김대현(DaeHyun Kim),조수정(Soo Jeong Cho) 한국생명과학회 2021 생명과학회지 Vol.31 No.2

본 연구에서는 우리나라 고유 수종인 노각나무(Stewartia koreana)의 부위별 이용 가능성을 알아보기 위하여 노각나무 잎과 가지를 70% 에탄올에 침지한 다음 부위별 항산화 활성을 조사하였다. 노각나무 가지 추출물(53.18±0.2 mg GAEs/extract g)에 비해 잎 추출물(162.57±0.9 mg GAEs/extract g)의 총 폴리페놀 함량이 높게 나타났고 플라보노이드 함량도 가지(4.7±0.1 mg QEs/extract g)에 비해 잎(59.1±0.9 mg QEs/extract g)에서 높게 나타났다. 노각나무 추출물의 DPPH 라디칼 소거능과 ABTS 라디칼 소거 활성은 잎과 가지 모두 농도 의존적으로 증가하였으며 가지보다는 잎에서 높게 나타났다. 노각나무 추출물 0.4 mg/ml의 농도에서 잎과 가지의 DPPH 라디칼 소거능은 각각 91.85±0.01%, 29.93±0.2%였으며 노각나무 잎 추출물은 양성대조구로 사용한 ascorbic acid (1mg/ml, 93.71±0.1%) 보다 낮은 농도에서 ascorbic acid와 유사한 DPPH 라디칼 소거능을 나타내었다. 노각나무추출물 0.4 mg/ml의 농도에서 잎과 가지의 ABTS 라디칼 소거능은 각각 94.77±0.34%, 25.13±0.15%였다. 노각나무잎과 가지의 ORAC 지수는 각각 104.13±0.48 μM TEs/ extract g과 57.36±0.35 μM TEs/extract g으로 가지보다는 잎의 ORAC 지수가 더 높게 나타났다. 노각나무 추출물의 세포 독성은 WST-1 assay를 이용하여 추출물이 섬유아세포 CCD-986sk에 미치는 영향으로 평가하였다. 이상의 결과를 종합해보면, 노각나무 잎 추출물은 가지에 비해 항산화 활성 지표인 총 폴리페놀과 플라보노이드 함량이 높고 항산화 활성이 높기 때문에 노각나무 잎 추출물은 항산화 활성이 우수한 기능성 식의약품소재나 화장품 소재로 사용될 수 있을 것으로 판단된다. This study was carried out to evaluate the antioxidant properties of the dried leaves and branches of Stewartia koreana Nakai. The dried leaf and branch of S. koreana were extracted with 70% ethanol at 80℃. The antioxidant activities of ethanol extracts of S. koreana leaf (EESL) and S. koreana branch (EESB) were analyzed. The total polyphenol contents in EESL and EESB were 162.57±0.9 mg of GAEs/extract g and 59.1±0.9 mg of GAEs/extract g, respectively. The flavonoid contents in EESL and EESB were 59.1±0.9 mg of QEs/extract g and 4.7±0.1 mg of QEs/extract g, respectively. EESL showed a better scavenging ability with DPPH and ABTS than EESB, at 0.4 mg/ml. Moreover, EESL were more effective according to ORAC values than EESL. The toxicity of EESL was investigated using a WST-1 assay on the human skin fibroblast cell line CCD-986sk. Therefore, EESL can be used as a potential source of functional, naturally-sourced material in cosmetics as well as food.

Porphyromonas gingivalis에 대한 노각나무 잎 추출물의 항균활성 및 생물막 형성 억제 효과

김혜수(Hye Soo Kim),박민정(Min Jeong Park),김수정(Soo Jeong Kim),김부경(Bu Kyung Kim),박준호(JunHo Park),김대현(DaeHyun Kim),조수정(Soo Jeong Cho) 한국생명과학회 2021 생명과학회지 Vol.31 No.3

본 연구에서는 천연물유래 구강건강 개선소재로써 노각나무의 이용 가능성을 알아보기 위해 노각나무 잎과 줄기를 에탄올에 추출한 다음 구강미생물에 대한 추출물의 항균활성을 조사하였다. 노각나무 잎과 줄기 추출물(1 mg/disc)은 구강미생물 중 P. gingivalis KCTC5352에 대해서만 항균활성을 나타내었으며 줄기보다는 잎 추출물의 항균활성이 우수하였다. 시판되고 있는 구강케어제품에 사용되고 있는 항균제와 노각나무 잎 추출물의 항균활성을 비교한 결과, P. gingivalis에 대한 노각나무 잎 추출물과 양성대조구로 사용한 triclosan의 항균활성은 유사하게 나타났으며. P. gingivalis에 대한 노각나무 잎 추출물의 MIC는 0.4 mg/ml이고 정균작용을 하였다. 노각나무 잎 추출물이 0.2-2.0 mg/ml 농도로 처리된 배양액에서 P. gingivalis KCTC5352의 생물막 형성과 세균 생육은 추출물의 농도가 증가할수록 농도의존적으로 억제되는 경향을 보였다. 또한 노각나무 잎 추출물(1 mg/ml) 처리가 P. gingivalis의 생물막 형성에 미치는 영향을 주사전자현미경으로 관찰한 결과에 의하면 추출물을 처리하지 않은 대조구는 추출물 처리구에 비해 P. gingivalis가 군집을 이루며 모여 있었고 세포 주변에서 생물막이 관찰되었지만 추출물을 처리한 처리구의 세포 주변에서는 생물막을 관찰할 수 없었다. qRT-PCR을 이용하여 생물막 형성 초기 과정에서 치면 부착에 필수적인 섬모(fimbriae)관련 mRNA 발현 양상을 0조사한 결과, 노각나무 잎 추출물이 0.2-2.0 mg/ml의 농도로 처리된 배양액에서 fimA와 mfa1 유전자 발현은 추출물의 농도가 높아질수록 농도의존적으로 억제되는 것을 확인할 수 있었다. 이상의 결과를 종합하면 노각나무 잎 추출물은 치주질환 원인균인 P. gingivalis에 대한 항균 활성과 생물막 형성 억제능이 우수하기 때문에 천연물유래 구강건강 개선소재로써 이용 가능성이 높을 것으로 판단된다. This study was conducted to investigate the potential of Stewartia koreana as oral healthcare materials. The antibacterial activity of ethanol extracts from leaves and branches of S. koreana against oral bacteria was confirmed. The leaf and branch extracts (1 mg/disc) showed antibacterial activity against P. gingivalis only among several tested oral bacteria. The leaf extracts showed higher antibacterial activity, with values similar to those of chlorhexidine, which was used as a positive control. The MIC of the leaf extract against P. gingivalis was 0.4 mg/ml and showed bacteriostatic action. The inhibitory effects of the extract on biofilm formation and on gene expression related to biofilm formation by P. gingivalis were determined by biofilm biomass staining, scanning electron microscopy (SEM), and qRT-PCR analysis. The biofilm production rate and cell growth of P. gingivalis in the cultures treated with 0.2-2.0 mg/ml of S. koreana leaf extracts were significantly decreased in a concentration-dependent manner. The inhibitory effect on the formation of P. gingivalis biofilms at concentrations of 1 mg/ml was confirmed by SEM. The qRT-PCR analysis showed concentration-dependent suppression of the fimA and fimB gene expression associated with fimbriae formation in the cultures treated with 0.2-2.0 mg/ml S. koreana leaf extract. These results support the conclusion that S. koreana leaf extracts can be used as oral healthcare materials derived from natural materials, as demonstrated by the antibacterial action and inhibition of biofilm formation of P. gingivalis.

노각나무 가지의 Dihydrochalcone 성분

김대근,안달래,이사임,양재헌,임종필,전훈,은재순,김학용,이재혁,박정숙 한국생약학회 2011 생약학회지 Vol.42 No.3

Three dihydrochalcone compounds were isolated from the twigs of Stewartia koreana (Theaceae) through repeated column chromatography. Their chemical structures were elucidated as phloridizin, 2’,4,4’-trihydroxy-6’-methoxydihydrochalcone and confusoside, respectively, by spectroscopic analysis. These compounds were isolated for the first time from this plant.

Antioxidant Flavonoids from the Twigs of Stewartia koreana

Lee, Sa-Im,Yang, Jae-Heon,Kim, Dae-Keun The Korean Society of Applied Pharmacology 2010 Biomolecules & Therapeutics(구 응용약물학회지) Vol.22 No.4

In the course of screening for antioxidant compounds by measuring the radical scavenging effect on 1,1-diphenyl- 2-picrylhydrazyl (DPPH), a total extract of the twigs of Stewartia koreana (Theaceae) was found to show potent antioxidant activity. Subsequent activity-guided fractionation of the methanolic extract led to the isolation of six phenolic compounds, ampelopsin (1), catechin (2), proanthocyanidin-A2 (3), fraxin (4), (2R, 3R)-taxifolin-3-${\beta}$-D-glucopyranoside (5), and (2S, 3S)-taxifolin-3-${\beta}$-D-glucopyranoside (6), as active principles. Their structures were elucidated by spectroscopic studies. Compounds 1-6 were isolated for the first time from this plant. Among them, three compounds 1-3 showed the significant antioxidative effects on DPPH, and riboflavin originated superoxide quenching activity. In riboflavin-nitrobluetetrazolium (NBT)-light system, compound 1 showed better superoxide quenching activity than vitamin C.

I-SSR 분석에 의한 노각나무 천연집단의 유전변이

양병훈(Byeung-Hoon Yang),한상돈(Sang-Don Han),구영본(Yeong-Bon Koo),박용구(Yong-Goo Park) 한국자원식물학회 2006 한국자원식물학회지 Vol.19 No.1

본 연구는 우리나라 특산수종이며 조경 및 원예적 가치가 높은 노각나무 유전변이를 조사하기 위해 6개 천연집단을 선발하여 DNA I-SSR 표지자를 사용, 유전다양성 및 유전구조를 조사하였다. 5개의 I-SSR primer(#813, 815, 818, 820, 823)에서 총 61개의 증폭산물을 관찰할 수 있었으며, 유전 다양성을 나타내는 P(Percentage of polymorphic loci)값과S.I.(Shannon s information Index)가 남쪽에 분포하는 오봉산(P=88.5%, S.I.=0.467), 금산(P=86.9%, S.I.=0.427), 바랑산(P=83.6%, S.I.=0.425)집단이 높았으며, 북쪽(내륙)에 분포하는 소백산(P=80.3%, S.I.=0.396), 지리산(P=77.1%, S.I.=0.368), 가야산(P=75.4%, S.I.=0.358)집단은 낮았다. 전체 유전변이 중 11.8%만이 집단간에 기인하는 것으로 나타났고, 나머지 88.2%는 집단내 개체간의 차이에서 기인하였다. 유전거리를 이용하여 UPGMA법에 의한 유집분석을 실시한 결과 지리적 분포에 대한 뚜렷한 경향은 나타나지 않았다. We investigated the genetic variation in Stewartia koreana Nakai by examining 61 I-SSR amplicons in 120 individuals distributed among six natural populations in Korea. The overall percentage of polymorphic I-SSR amplicons was 81.9% and mean number of amplicons per I-SSR primer was 12.2. Levels of genetic diversity within 6 populations were similar each other[Shannon s Index 0.358~0.467(mean: 0.407)]. The Mt. Obong population had the highest level of genetic diversity and was most distinctive from the other populations. Most variation existed among individuals within population(88.2%). Genetic differentiation among populations(ΦST) was 0.118. The UPGMA dendrogram based on the genetic distance failed in showing decisive geographic relationships.