Publications

Detailed Information

Anti-inflammatory Mechanism of a Diterpenoid, Ebractenoid F, Isolated from Euphorbia ebracteolata Hayata

DC Field Value Language
dc.contributor.advisor김영식-
dc.contributor.author마상연-
dc.date.accessioned2018-05-29T04:41:44Z-
dc.date.available2019-04-17-
dc.date.issued2018-02-
dc.identifier.other000000149756-
dc.identifier.urihttps://hdl.handle.net/10371/142215-
dc.description학위논문 (석사)-- 서울대학교 대학원 : 약학대학 약학과, 2018. 2. 김영식.-
dc.description.abstractEuphorbia ebracteolata Hayata, belongs to Euphorbiaceae, is widely distributed through the south of China. In the traditional chinese medicine (TCM), the roots of E. ebracteolata have been prescribed for chronic inflammation-mediated diseases such as tracheitis, cutaneous tuberculosis, tumor, and psoriasis. It has been reported that E. ebracteolata is rich in bioactive components such as acetophenones, flavonoids, and diterpenes. Among the constituents of E. ebracteolata, diterpenes in E. ebracteolata have effects on anti-inflammation, anti-tumor, and anti-fungal activities. However, the anti-inflammatory mechanism of most of constituents in E. ebracteolata Radix is not yet fully discovered.
For find the anti-inflammatory compound from E. ebracteolata Radix, bioassay-guided fractionation, which is a general way to isolate and characterize bioactive compounds from natural products, was conducted. It might be a worth way to find out the most potential compounds possessing biological activities. Thus, we applied this concept to process separation. In addition, a separation method, called high speed counter-current chromatography (HSCCC), was co-operated because it provides us many advantages in terms of high yield, sample loading capacity, easy scale-up, purity, and resolution. Following the bioassay-guided isolation, the compounds were elucidated and identified by various spectroscopic ways. Ebractenoid F (EF) showing the potential effects was ultimately obtained by this method, bioassay-guided isolation.
Inflammation is a key factor of protection system triggered by immune cells in response to harmful stimuli. It mediates various cellular processes such as proliferation, cell cycle, and differentiation. If the stimuli remain, the acute inflammatory response is connected to the chronic inflammatory response. The nitric oxide (NO) and SEAP are the inflammatory mediators, so they could arise from the inflammation. In this vein of thought, we seek for anti-inflammatory agents which can reverse both NO and SEAP production regulated by NF-κB.
EF shows the prominent inhibition of NF-κB Secretary Alkaline Phosphatase (SEAP) (IC50 = 7.71 μM), implying that it is specifically targeting on NF-κB. It decreases the downstream of NF-κB including various pro-inflammatory mediators, such as inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and interleukin-6 (IL-6), at both protein and mRNA levels. Moreover, it down-regulates phosphorylation and degradation of inhibitory κB (IκB)-α in 20 minutes. Nuclear translocation and transcriptional activity are suppressed when cells are treated with EF. In addition, we investigated whether it has an effect on LPS-induced mitogen-activated protein kinase (MAPKs, p-ERK, and p-JNK) and the upstream signaling pathways (p-AKT and p-IKK).
Taken together, we tried to find the anti-inflammatory compound from E. ebracteolata using bioassay-guided fractionation in that there are not enough studies. At this research, we could discover a potential compound, ebractenoid F (EF). Through this achievement, we also expected that inflammation-mediated diseases can be managed with this compound since the NF-κB signaling pathway related to inflammation was suppressed. Although we still need to study on the effect of ebractenoid F, it is possible to be a key regulator on inflammation-related to diseases.
-
dc.description.tableofcontentsⅠ. INTRODUCTION 1
1. Bioassay-guided Isolation 1
2. Inflammation 3
3. Euphorbia ebracteolata Hayata 6
4. NF-κB 7
5. High Speed Counter-Current Chromatography (HSCCC) 9
6. Purpose of This Study 12
Ⅱ. MATERIALS & METHODS 13
1. MATERIALS 13
1.1 Plant Materials 13
1.2 Chemicals and Reagents 13
1.3 Instruments 14
2. METHODS 16
2.1 Solvent extraction of Euphorbia ebracteolata Hayata 16
2.2 Sub-fractionation of Euphorbia ebracteolata Hayata using High Speed Counter Current Chromatography (HSCCC) 16
2.3 Isolation of an anti-inflammatory compound via preparative-HPLC 17
2.4 HPLC analysis 17
2.5 Identification of isolated compounds 18
2.6 Cell Culture 18
2.7 Cell Viability assay 19
2.8 The Measurement of Nitric Oxide (NO) Production 20
2.9 NF-κB SEAP reporter gene assay 20
2.10 NF-κB Luciferase assay 21
2.11 Western Blot analysis 21
2.12 Quantitative real-time reverse transcriptase polymerase chain reaction (PCR) 22
2.13 Preparation of cytoplasmic and nuclear extract 25
2.14 Statistical analysis 25
Ⅲ. RESULTS 26
1. Isolation of Ebractenoid F (EF) from E. ebracteolata by bioassay-guided fractionation 26
2. Screening Extract, Fractions, Sub-fractions, and Compounds from E. ebracteolata 31
3. Elucidation and Determination of Ebractenoid F (EF) 35
4. Effects of Ebractenoid F (EF) on pro-inflammatory mediators in LPS-stimulated RAW 264.7 cells 44
5. Effects of Ebractenoid F (EF) on LPS-mediated NF-κB trans-locational and transcriptional activity 47
6. Effects of Ebractenoid F (EF) on the upstream of NF-κB and MAPKs pathway 49
Ⅳ. DISCUSSION 52
Ⅴ. REFERENCES 56
ABSTRACT IN KOREAN 62
-
dc.formatapplication/pdf-
dc.format.extent2616888 bytes-
dc.format.mediumapplication/pdf-
dc.language.isoen-
dc.publisher서울대학교 대학원-
dc.subjectEuphorbia ebracteolata Hayata-
dc.subjectHSCCC-
dc.subjectNF-κB-
dc.subjectRAW 264.7 cell-
dc.subject.ddc615-
dc.titleAnti-inflammatory Mechanism of a Diterpenoid, Ebractenoid F, Isolated from Euphorbia ebracteolata Hayata-
dc.typeThesis-
dc.description.degreeMaster-
dc.contributor.affiliation약학대학 약학과-
dc.date.awarded2018-02-
Appears in Collections:
Files in This Item:

Altmetrics

Item View & Download Count

  • mendeley

Items in S-Space are protected by copyright, with all rights reserved, unless otherwise indicated.

Share