[보고서]재래귤 유래 항염증 천연활성물질 발굴 및 제품화에 관한 연구

재래귤 유래 항염증 천연활성물질 발굴 및 제품화에 관한 연구
Investigation and commercialization of anti-inflammatory phytochemicals originated from Jeju traditional Citrus 원문보기

보고서 정보
주관연구기관	제주대학교 Cheju National University
보고서유형	최종보고서
발행국가	대한민국
언어	한국어
발행년월	2014-03
과제시작연도	2011
주관부처	농림축산식품부 Ministry of Agriculture, Food and Rural Affairs(MAFRA)
등록번호	TRKO201400005701
과제고유번호	1545003296
사업명	생명산업기술개발
DB 구축일자	2014-11-29
DOI	https://doi.org/10.23000/TRKO201400005701

초록 ▼

Ⅳ. 연구개발결과
○ 재래감귤 성숙과 추출물의 세포변이억제 효능 및 성분분석
- 주원료 선정을 위한 성숙과 추출물의 암세포증식 억제 및 암세포사멸 효능을 평가했으며 열수추출물과 80% 에탄올추출 방법에 의한 제주 재래감귤 추출물은 암세포에 대한 항암효능이 크게 나타나지 않는다는 것을 확인함.
- 이예감, 당유자, 팔삭 초임계 추출물을 확보하고 GC/MS를 통한 재래 감귤 초임계 추출물 성분을 분석함.
○ 재래감귤 미성숙과 초임계 추출물의 세포변이억제 효능 및 성분분석
- 미성숙 재래감귤의 열수, 80% 에탄올 추출물들에 대한 세포증식 억제 효능 평가
- 미성숙 이예감, 당유자, 팔삭 초임계 추출물을 확보하고, GC/MS를 통한 재래 감귤 초임계 추출물 성분을 분석함.
○ 재래귤의 다양한 추출물에 대한 대식세포의 세포독성 측정
- 재래귤의 다양한 추출물에 대한 대식세포의 세포독성 측정하고 재래귤의 과피 추출물에 대한 LPS로 유도된 산화질소 억제능을 확인함.
○ 구아바 잎 분획물의 항산화효능 탐색
- 구아바 잎의 다양한 용매 추출물을 확보하고 구아바 잎 분획물들에 대한 항산화 효능 비교 탐색
- 산화 스트레스에 의해 유도된 DNA damage에 대한 구아바 잎 분획물들의 세포 보호 효능을 확인함
○ 인진쑥으로부터 분리된 Capillarisin의 항염증 효능 탐색 및 항암 기전 구명
- 동물세포계 (in vitro)를 이용한 Capillarisin의 항염증효능을 평가하고 apoptosis 유발에 의한 암세포사멸 기작을 확임함.
○ 당유자 과피 에션셜 오일에 의한 U373MG 세포에서의 세포 증식 억제 및 세포사멸 효능 평가
- 당유자 과피 에션셜 오일에 의한 U373MG 세포에서의 세포 증식 억제 효능 탐색과 당유자 과피 에션셜 오일 성분을 GC/MS로 분석함.
○ 당유자 잎 분획물의 세포변이억제효능 탐색 및 활성에 기여하는 천연활성물질(phytochemicals) 발굴
- 당유자 잎에서 분리한 여러가지 분획물에 대한 전립선 암세포 세포사멸기작 확인하고 Nobiletin에 의한 전립선 암세포 constitutive STAT3 활성도 저해능 평가함.
- Nobiletin에 의한 SNU-16 위암 세포에서의 apoptosis현상이 ER-stress 유발과 관련된 신호 전달에 기인한 것임을 밝힘.
○ 이예감 과피 초임계 추출물의 세포성장 억제 및 apoptosis 활성
- 이예감 과피 추출물 (supercritical extracts of yeagam peel (SEYG))에 대한 다양한 암세포에서의 세포독성 측정하고 암세포에서의 constitutive STAT3 활성도 저해능 평가 및 DU145 이종이식 마우스 모델을 이용한 생체내 (in vivo) 실험을 통해 항암 활성을 확인함.
○ 팔삭 추출물의 항전이작용 및 기전규명
- 팔삭 과피 추출물 (supercritical extracts of phalsak peel (SEPS))에 대한 다양한 암세 포에서의 CXCR4 측정하고, SEPS에 의한 암세포의 항전이작용 기전을 확인함.
- 팔삭 과피 추출물에서의 유효성분으로 예상되는 osthole과 auraptene에 의한 암세포 사멸 효능을 확인함.
○ 팔삭과피와 구아바의 최적혼합물로부터 항염증 활성 및 기전연구
- 동물세포계(in vitro)를 이용한 OM의 항염증효능
• RAW264.7 대식세포에서 lipopolysaccharide (LPS)로 산화질소의 발생을 유도하였을 때 LPS로 유도된 산화질소의 생성이 혼합물 농도 의존적으로 억제되는 것을 확인함
- 동물 (in vivo)을 이용한 OM의 항염증효능
• 동물모델에서 LPS에 의한 충격으로부터 혼합물의 보호 효과를 평가하기 위해 우리는 치명적인 내독소 혈증과 마우스의 사망률에 미치는 영향을 모니터링.
○ 천연물 혼합제재를 이용한 제품 개발
- 예비실험을 통해 우수한 항염증 활성을 나타내는 구아바와 인진호를 최종 부원료로 선정하였으며 추출조건에 따른 추출특성의 모니터링 및 추출조건의 최적화를 위하여 반응표면분석법 (response surface methodology, RSM)을 이용하여 구아바 잎 및 인진호의 추출조건의 최적화를 확립.
- 항염증 활성이 우수한 기능성음료를 위한 재래감귤 팔삭 및 구아바, 인진호의 배합비 최적화
- 팔삭-구아바 혼합제재의 정제 타정용 시료 조제 및 구성약물의 품질표준화
- 주원료의 구성약물의 품질표준화
- 공정별 품질관리에 대한 항염증 활성 및 위생안전성 측정

Abstract ▼

IV. Results
○ Anti-proliferation and apoptotic activity and active material analysis of Jeju traditional Citrus (mature fruit) Extracts
- Screening anti-proliferation and apoptotic activity of various mature citrus extracts and water extracts and 80% ethanol extracts show no apoptosis activity.
- Prepare supercritical extracts of mature Iyegam, dangyuja, palsack and analyse supercritical extracts using GC/MS.
○ Anti-proliferation and apoptotic activity and active material analysis of Jeju traditional Citrus (immature fruit) Extracts
- Screening anti-proliferation and apoptotic activity of various immature citrus extracts and water extracts and 80% ethanol extracts show no apoptosis activity.
- Prepare supercritical extracts of immature Iyegam, dangyuja, palsack and analyse supercritical extracts using GC/MS.
○ Measure cytotoxicity of extracts of Jeju traditional citrus on macrophage cells
- Measure cytotoxicity of extracts of Jeju traditional citrus on macrophage cells and find inhibition of Nitric oxide generation of peel extract from traditional citrus.
○ Screening the anti-oxidation activity of extracts from leaf of Guava
- Obtain various guava leaf extracts and Screening the anti-oxidation activity of them
- Conform the protective effects of various fraction from leaf of Guava against oxidative DNA damage.
○ Screening and identification of anti-inflammation and anti cancer activity of capillaricin from capillais
- Evaluate anti-inflammatory activity of capillaricin in vitro assay and elucidate mechanism of apoptosis
○ Evaluate anti-proliferation effects of essential oil from dangyuja peel against U373MG cells
- Evaluate anti-proliferation effects of essential oil from dangyuja peel against U373MG cells and analyse essential oil from dangyuja peel using GC/MS.
○ Screen dangyuja leaf extracts for the inhibitory activity of trans-differentiation of cancer cells and identify the phytochemical candidate of action.
- conform dangyuja leaf extracts for the apoptosis activity against prostate cancer cell line and show nobiletin inhibitory activity on constitutive STAT3.
- Show that apoptosis of SNU-16 stomach cancer cells induced by nobiletin is due to signaling via ER-stress.
○ Anti-proliferative activity of supercritical extracts of yeagam peel (SEYG)
- Measure the anti-proliferative activity of supercritical extracts of yeagam peel (SEYG) against various cancer cells and show SEYG inhibit constitutive STAT3.SEYG also anti-proliferative activity in vivo DU145 tumour mouse model.
- Measure the anti-proliferative activity of osthole and auraptene from supercritical extracts of yeagam peel (SEYG) against stomach cancer cells and show osthole and auraptene induce apoptosis.
○ Migration inhibitory effect of (supercritical extracts of phalsak peel (SEPS)) and mechanism
- Measure the CXCR4 in SEPS treated cancer cells and show inhibited migration of cancer cells.
○ Mechanism study of anti-inflammatory effects of optimum mixture of SEPS and danyuja(OM)
- Anti-inflammation assay of OM in vitro
• LPS induced NO was reduced by MO treatment in RAW264.7 macrophage cells
- Anti-inflammation assay of OM in vivo
• In animal model, measure protection effects of OM against LPS endotoxin and mouse death.
○ Products development using phytochemical mixture
- Select Guava and capillaris as final additive for anti-inflammatory biomaterial. Using response surface methodology, RSM, optimize the extraction condition.
- Select best ratio of pasak, Guava and capillaris for anti-inflammation activity
- Standardization of composition of mixture and produce tablet using extract mixture of Pasak and Guava
- Standardization of composition of main source

목차 Contents

표지 ... 1
제출문 ... 2
요 약 문 ... 3
SUMMARY ... 6
CONTENTS ... 9
목차 ... 10
제 1 장 연구개발과제의 개요 ... 13
제 1절 연구개발의 필요성 ... 13
제 2절 연구개발의 목표 및 범위 ... 15
제 2 장 국내외 기술개발 현황 ... 18
1. 관련기술현황 ... 18
2. 국내연구개발의 필요성 ... 19
제 3 장 연구개발수행 내용 및 결과 ... 21
제 1절 재래감귤 성숙과 추출물의 세포변이억제 효능 및 성분분석 ... 21
제 2절 재래감귤 미성숙과 추출물의 세포변이억제 효능 및 성분 분석 ... 47
제 3절 재래감귤의 다양한 추출물에 대한 대식세포의 세포독성 측정 ... 61
제 4절 부재료로 사용된 구아바 잎 EtOAc 분획물과 인진쑥 추출물에서 분리된 Capillarisin에 의한 생리활성기작 탐색 ... 66
제 5절 당유자 과피 에션셜 오일에 의한 U373MG 세포에서의 세포 증식 억제 및 세포사멸 효능 평가 ... 80
제 6절 당유자 잎 분획물의 세포변이억제효능 탐색 및 활성에 기여하는 천연활성물질(phytochemicals) 발굴 ... 83
제 7절 이예감 과피 초임계 추출물의 STAT3 활성저해 및 세포성장 억제 ... 93
제 8절 팔삭 추출물의 항전이작용 및 기전 구명 ... 103
제 9절 팔삭과피와 구아바의 최적혼합물로부터 항염증 활성 및 기전 연구 ... 111
제 10절 천연물 혼합제재를 이용한 제품 개발 ... 116
제 4 장 목표달성도 및 관련분야에의 기여도 ... 177
제 1절 평가의 착안점 및 달성도 ... 177
제 2절 관련분야의 기여도 ... 180
제 5 장 연구개발결과의 활용계획 ... 181
제 1절 실용화·산업화 계획 ... 181
제 2절 추가연구개발 ... 182
제 6 장 연구개발과정에서 수집한 해외과학기술정보 ... 183
제 1절 국내 제품생산 및 시장 현황 ... 183
제 2절 국외 제품생산 및 시장 현황 ... 183
제 3절 파이토케미컬 유래 항암소재 개발 현황 ... 184
제 7 장 참고문헌 ... 185
끝페이지 ... 187

표/그림 (182)

표 대표적인 재래귤
표 Yield of Hot water and 80% ethanol extract.
표 Yield of CO2 supercritical extract.
표 Jeju traditional citrus super critical extract inhibits the cell proliferation on AGS(left) and HeLa(right) cell lines.
표 Effect of Jeju traditional citrus super critical extract on cell morphology.
표 Effect of Jeju traditional citrus super critical extract on cell morphology.
표 FACS analysis of Sub G1 phase (%) in the HeLa cells treated with extract.
표 FACS analysis of Sub G1 phase (%) in the AGS cells treated with extract.
표 Jeju traditional mature Dangyuja super critical extract inhibits the cell
표 Effect of Jeju traditional mature Dangyuja peel super critical extract on cell morphology
표 Analysis of % of Sub G1 phase in the HeLa cells treated with peel extract.
표 DNA fragmentation of HeLa cells induced by Jeju traditional mature Dangyuja peel super critical extract (Q50: Quercetin 50 μM).
표 Western blot analysis of apoptosis-related protein expression in mature Dangyuja peel super critical extract-treated HeLa cells (Q50: Quercetin 50 μM).
표 Supercritical CO extract from palsak induces cell death in breast cancer stem2 cells.
표 Supercritical CO extract from palsak induces apoptotic cell death in breast2 cancer stem cells.
표 Supercritical CO extract from palsak down regulate multi-drug resistance gene2 ABCG2 in breast cancer stem cells.
표 GC-MS chromatogram of supercritical extract of Y:Yagam varieties from Jeju,Korea.
표 Relative content of functional groups of volatile organic compounds identified in supercritical extract of Y:Yegam.
표 Relative number of compounds belongs to functional groups identified in supercritical extract of Y:Yegam.
표 Relative proportion of major five constituents in supercritical extract Yegam fruit.
표 Relative proportion of compounds those found in supercritical extract of Yegam citrus fruit but not in other two samples.
표 GC-MS chromatogram of supercritical extract of Dangyuja varieties from Jeju,Korea.
표 Relative content of functional groups identified from Dangyuja fruits.
표 Relative content of number of compounds based upon the functional groups identified from Dangyuja fruits.
표 Relative proportion of major five constituents in supercritical extract of Dangyuja fruit.
표 Relative proportion of constituents those found in supercritical extract of Dangyuja fruit but not in other two varieties.
표 GC-MS chromatogram of supercritical extract of Palsak fruit from Jeju Korea.
표 Relative content of functional groups identified from supercritical extract of Palsak.
표 Relative content of number of compounds based upon the functional groups in Palsak fruit.
표 Relative proportion of major five constituents in supercritical extract of Palsak fruit.
표 Relative proportion of compounds those found in supercritical extract of Palsak fruit but not in other two varieties.
표 Comparative profile of functional groups among the fruit samples.
표 Yield of Hot water and 80% ethanol extract.
표 Jeju traditional immature citrus peel 80% ethanol extracts inhibits the cell proliferation on AGS and HeLa cell lines.
표 Yield of Hot water and 80% ethanol extract.
표 Jeju traditional immature citrus super critical extract inhibits the cell
표 Effect of Jeju traditional immature citrus super critical extract on cell morphology
표 Effect of Jeju traditional immature citrus super critical extract on cell morphology
표 Analysis of % of Sub G1 phase in the HeLa cells treated with peel extracts.
표 Analysis of % of Sub G1 phase in the AGS cells treated with peel extracts.
표 GC-MS chromatogram of supercritical extract of immature dangyuja varieties from Jeju,Korea.
표 Relative content of functional groups of volatile organic compounds identified in supercritical extract of immature Dangyuja.
표 Relative proportion of major four constituents in supercritical extract of immature Dangyuja.
표 GC-MS chromatogram of supercritical extract of immature Phalsak varieties from Jeju, Korea.
표 Relative content of functional groups of volatile organic compounds identified in supercritical extract of immature Phalsak.
표 Relative proportion of major four constituents in supercritical extract of immature Phalsak.
표 GC-MS chromatogram of supercritical extract of immature Yeagam varieties from Jeju, Korea.
표 Relative content of functional groups of volatile organic compounds identified in supercritical extract of immature Yeagam.
표 Relative proportion of major five constituents in supercritical extract of immature Yeagam.
$Effects of fraction from various Citrus on the cell viability in RAW 264.7 macrophages.$ 표 Effects of fraction from various Citrus on the cell viability in RAW 264.7 macrophages.
$Effects of fraction from various Citrus on the LPS‐induced nitric oxide (NO) in RAW 264.7 macrophages.$ 표 Effects of fraction from various Citrus on the LPS‐induced nitric oxide (NO) in RAW 264.7 macrophages.
$Effects of fraction from Iyokan and hassaku pericarp on the LPS‐induced nitric oxide (NO) in RAW 264.7 macrophages.$ 표 Effects of fraction from Iyokan and hassaku pericarp on the LPS‐induced nitric oxide (NO) in RAW 264.7 macrophages.
$Inhibition of the fraction from Iyokan and hassaku pericarp on LPS‐induced iNOS,COX-2 gene products in RAW 264.7 macrophages (A) 이예감 과피. (B) 팔삭 과피.$ 표 Inhibition of the fraction from Iyokan and hassaku pericarp on LPS‐induced iNOS,COX-2 gene products in RAW 264.7 macrophages (A) 이예감 과피. (B) 팔삭 과피.
$Inhibition of iNOS and COX-2 mRNA expressions by fraction from Iyokan and hassaku pericarp in LPS-stimulated RAW264.7 cells (A) 이예감 과피. (B) 팔삭 과피.$ 표 Inhibition of iNOS and COX-2 mRNA expressions by fraction from Iyokan and hassaku pericarp in LPS-stimulated RAW264.7 cells (A) 이예감 과피. (B) 팔삭 과피.
$Suppression of LPS‐induced NF-κB activity by fraction from Citrus hassaku pericarp in RAW 264.7 macrophages.$ 표 Suppression of LPS‐induced NF-κB activity by fraction from Citrus hassaku pericarp in RAW 264.7 macrophages.
$Effects of fraction from Citrus hassaku pericarp on LPS‐induced activation of p38, ERK, and JNK.$ 표 Effects of fraction from Citrus hassaku pericarp on LPS‐induced activation of p38, ERK, and JNK.
$Inhibition of the fraction from Iyokan and hassaku pericarp on LPS‐induced IL-6 and TNFa production in RAW 264.7 macrophages.$ 표 Inhibition of the fraction from Iyokan and hassaku pericarp on LPS‐induced IL-6 and TNFa production in RAW 264.7 macrophages.
$Total phenolic content and total flavonoid content from different fraction of P. cattleianum leaf extract.$ 표 Total phenolic content and total flavonoid content from different fraction of P. cattleianum leaf extract.
$Radical scavenging activity of various fractions of P. cattleianum leaf extract.$ 표 Radical scavenging activity of various fractions of P. cattleianum leaf extract.
표 Protective effects of EAF of P. cattleianum leaf.
표 Protective effects of EAF on H O -induced DNA damage by comet assay.
표 (A) Cells were pretreated with different concentrations of CPS for 1 h and stimulated with LPS (1 ug/mL) for 24 h. The values obtained were compared with those of standard concentrations of sodium nitrite dissolved in Dulbecco's modified Eagle's minimal essential medium (DMEM), and the concentrations of nitrite in a conditioned media of sample‐treated cells were calculated. Data were obtained from three independent experiments and were expressed as means ± SD.
표 (A) Cells were pretreated with 160 uM of CPS for 1 h and stimulated with LPS (1 ug/mL) for the indicated times and the nuclear extracts were prepared, resolved by SDS-PAGE, and electrotransferred to a PVDF membrane, after which Western blot analyses using anti‐p65 antibody was performed. Lamin B protein was used as a loading control. The results shown are representative of the three independent experiments. (B) Cells were pretreated with the indicated concentrations of CPS for 1 h and stimulated with LPS (1 ug/mL) for 1 h; nuclear extracts were prepared as described in the methods section. Detection of NF-kB binding activities was performed by electrophoretic mobility shift assay (EMSA). The specificity of NF-kB complex formation was verified in the LPS ‐only sample by displacement with a 50-fold excess of the unlabeled mutated oligonucleotide (mut) and a 50-fold excess of the unlabeled consensus oligonucleotide (cold) (Data not shown).
표 (A, B, and C) The cells were pretreated with the indicated concentrations of CPS for 1 h and stimulated with LPS (1 ug/mL) for 1 h; whole‐cell extracts were prepared, resolved by SDS-PAGE, and electrotransferred to a PVDF membrane. After which Western blot analysis using phospho-specific anti-ERK, phospho-specific anti-JNK, and phospho-specific anti-p38 antibodies was performed. The same membranes were reblotted with anti-ERK, anti-JNK, and anti-p38 antibodies. The results shown are representative of the three independent experiments.
표 Cells were pretreated with different concentrations of CPS for 1 h and stimulated with LPS (1 ug/mL final concentration) for 24 h. (A) The amount of TNF-α release was determined by a TNF-α antibody‐coated enzyme‐linked immunosorbent assay (ELISA) kit, as described in the Materials and methods. (B) The amount of IL-6 release was determined by an IL-6 antibody coated ELISA kit, as described in the Materials and Methods. (C) The amount of PGE2 release was determined by the mouse PGE2 ELISA kit. Data were obtained from three independent experiments and were expressed as means ± SD. (D) The amount of IL-1β release was determined by an IL-1b antibody‐coated ELISA kit.
표 (A) CPS suppresses cell proliferation in U266 cells were plated in triplicate, treated with 50, 100, and 150 uM CPS, and then subjected to MTT assay after 12, 24, and 36 h to analyze proliferation of cells. (B) U266 cells were treated 150 uM of CPS for 24 h, after which the cells were washed, fixed, stained with PI, and analyzed for DNA content by flow cytometry.
표 U266 cells (1 × 106 cells/mL) were treated with 0, 30, 50, 100, and 150 uM CPS for 6 h.
표 After 6 h of CPS treatment, the cells were fixed and permeabilized.
표 (A) U266 cells were treated with the indicated concentrations of pervanadateand 150 uM CPS for 6 h. Whole-cell extracts were prepared and immunoblotted with antibodies for phospho-STAT3 (Tyr705). The same blots were stripped and reprobed with STAT3 antibody to verify equal protein loading. (B and C) U266 cells were treated with the indicated concentrations of CPS for 6 h. (B) Whole-cell extracts were prepared and immunoblotted with antibodies for SHP-1, SHP-2, and PTEN. The same blots were stripped and reprobed with β-actin antibody to verify equal protein loading. (C) Total RNA was extracted and examined for expression of SHP-1, SHP-2, and PTEN by Reverse Transcription-Polymerase Chain Reaction (RT-PCR). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as an internal control to show equal RNA loading.
표 (A and B) U266 cells were treated 150 uM of CPS for 24 h, incubated with an anti-Annexin V antibody conjugated with fluorescein isothiocyanate (A), TUNEL-Label mix conjugated with the TUNEL-Enzyme (B) and then analyzed with a flow cytometer for apoptotic DNA fragmentation effects. (C and D) U266 cells were treated with the indicated concentrations of CPS for 24 h. (C) Whole-cell extracts were prepared; 30 ug portions of those extracts were resolved on 10% SDS-PAGE and probed against Bcl-xl, Bcl-2, Cyclin D1, Survivin, and IAP1 antibodies. The same blots were stripped and reprobed with β -actin antibody to verify equal protein loading. (D) Total RNA was extracted and examined for the expression of Bcl-xl, Bcl-2, CyclinD1, and Survivin by RT-PCR. GAPDH was used as an internal control to show equal RNA loading.
표 (A) U266 cells were treated with the indicated concentrations of CPS for 24 h. Whole-cell extracts were prepared and immuneblotted with antibodies for Caspase-3 and PARP. The same blots were stripped and reprobed with β-actin antibody to verify equal protein loading. (B) U266 cells were treated with the indicated concentrations of CPS for 6 h and analyzed for nuclear STAT3 levels by EMSA (left panel) . U266 cells (1 × 106 cells/mL) were treated with 150 uM CPS for the indicated times and analyzed for nuclear STAT3 levels by EMSA (right panel) .
표 U266 cells (1 × 106 cells/well) were cotreated with CPS for 6 h prior to bortezomib treatment and added with bortezomib for 24 h, (A) And then whole-cell extracts were prepared; 30 ug portions of those extracts were resolved on 10% SDS-PAGE and probed against anti-apoptotic, proliferative, and metastatic, and angiogenic gene products. The same blots were stripped and reprobed with β-actin antibody to verify equal protein loading. (B) Whole-cell extracts were prepared and immunoblotted with antibodies against Caspase-3 and PARP. The same blots were stripped and reprobed with β-actin antibody to verify equal protein loading.
표 Cell growth inhibition.
표 Effects of DPEO on nuclear Hoechst33342 staining and DNA fragmentation.
표 The percentage of cell cycle phase in the DPEO-treated U373 cells for 12 h.
표 Effects of DPEO on mitochondria membrane potential.
표 Immunoblot analysis of apoptosis-related protein expression in DPEO treated U373 cells.
표 Major components (in %) of Dangyuja peel essential oil separated by gas chromatography-mass spectroscopy.
$Effect of the chloroform (CHCl3) fraction of Dangyuja leaves (DCF) on cytotoxicity.$ 표 Effect of the chloroform (CHCl3) fraction of Dangyuja leaves (DCF) on cytotoxicity.
표 Effect of DCF on proliferation and exerts strong cytotoxicity against DU145 cells compared with noncancerous RWPE-1 human prostate cells.
표 Effect of DCF on cell cycle.
표 Effect of DCF on cell cycle.
표 DCF suppresses constitutive activation of Janus like kinase (JAK) 2.
표 DCF suppresses constitutive activation Src.
표 AG490 suppresses cell proliferation and indeces caspase-3 activation.
표 Identification of nobiletin in DCF.
표 Nobiletin suppresses constitutive activation of STAT3, JAK and Src.
표 Nobiletin induces apoptosis by PARP cleavage.
표 Nobiletin induces apoptosis by PARP cleavage.
표 Proteins identified from the 2-DE gels of human gastric cancer SNU-16 cell line by PMF.
표 RT-PCR and Western blot analysis of protein expression in gastric cancer cells.
표 Expression level of cell growth and ER-stress related proteins in SNU-16 cells.
표 Chloroquine inhibits autophagy and sensitizes SNU-16 cells to cytotoxic actions of nobiletin.
표 Inhibition of autophagy enhance nobiletin-induced apoptosis.
표 U266, K562, DU145, MDA-MB-231, and HepG2 cells were incubated at 37 ℃ with various indicated concentrations of SEYG for 24 h, and the viable cells were assayed using the MTT reagent.
표 Cells were incubated with the indicated concentrations of SEYG for 6 h.
표 DU145 cells (1 × 106 cells/well) were treated with various indicated concentrations of SEYG and time intervals.
표 SEYG suppresses STAT3 binding activity.
표 SEYG causes the inhibition of translocation of STAT3 to the nucleus.
표 Cell proliferation assay was performed using the Roche xCELLigence Real-Time Cell Analyzer (RTCA) DP instrument (Roche Diagnostics GmbH, Germany). After DU145 cells were seeded onto 96-well E-plates and continuously monitored using impedance technology.
표 DU145 cells were incubated with the indicated concentrations of SEYG for 24 h.
표 (A) After DU145 cells were seeded onto 6-well plates, they were incubated with the indicated concentrations of SEYG for 24 h. Then, the cells were fixed and analyzed using a flow cytometry. (B) DU145 cells were incubated with the indicated concentrations of SEYG for 24 h.
표 (A) Cells were treated with indicated concentrations of SEYG for 24 h. Afterward, they were incubated with anti-annexin V antibody conjugated with FITC plus PI and analyzed with a flow cytometer for apoptotic effects. (B) Cells were treated with indicated concentrations of SEYG for 24 h. Cells were fixed, stained with TUNEL assay reagent, and then analyzed with flow cytometer.
표 After DU145 cells were seeded onto 6-well plates, they were treated with various indicated concentrations of SEYG for 24 h.
표 (A) Schematic representation of experimental protocol described in “Materials and Methods.” DU145 cells (4 × 106 cells/mice) were injected subcutaneously into the right flank of the mice. The animals were randomized after 1 week of tumor cell injection into four groups based on tumor volume. Group I was given PBS (200 μL, i.p. thrice/week), group II was given SEYG (50 mg/kg body weight, i.p. thrice/week), and group III was given SEYG (200 mg/kg body weight, i.p. thrice/week). (B) Necropsy photographs of mice bearing subcutaneously implanted prostate tumors. (C) Tumor volumes in mice measured during the course of experiment and calculated using the formula V = 4 / 3 πr3,** indicates P < 0.01. (D) Tumor volumes in mice measured on the last day of the experiment at autopsy using Vernier calipers and calculated using the formula V = 4 / 3 π r3 (n=11). Columns, mean; bars, SE.
표 (A) Immunohistochemical analysis of proliferation marker Ki-67+ cell indicates the inhibition of human prostate cancer cells proliferation by SEYG dose-dependent treated groups of animals. Samples from 3 animals in each treatment group were analyzed. (B) Immunohistochemical analysis of CD31 for microvessel density in prostate tumors indicates the inhibition of angiogenesis by SEYG dose-dependent treated groups of animals. Samples from 3 animals in each treatment group were analyzed. (C) Immunohistochemical analysis of cleaved caspase-3 in prostate tumors. Samples from 3 animals in each treatment group were analyzed.
표 Western blot analysis showed the inhibition of p-STAT3, p-JAK1, p-JAK2, and p-Src by SEYG in whole cell extracts from animal tissue. The same blots were stripped and reprobed with STAT3, JAK1, JAK2, and Src antibody to verify equal protein loading.
표 Equal amounts of lysates were analyzed by Western blot analysis using antibodies against bcl-2, bcl-xL, survivin, IAP-1, IAP-2 (A), COX-2, cyclin D1, VEGF, and MMP-9 (B), p53, and p21 (C). β-actin was used as a loading control. Western blotting samples from three mice in each group were analyzed and representative data are shown.
표 U266, K562, DU145, MDA-MB-231, and HepG2 cells were incubated at 37 ℃ with 100 ug/mL of SEPS for 24 h.
표 Cells were incubated with the indicated concentrations of SEPS for 24 h.
표 (A) MDA-MB-231 cells were treated with various indicated concentrations of SEPS. Then equal amounts of lysates were analyzed by Western blot analysis using antibody against CXCR4. (B) MDA-MB-231 cells were treated with various indicated time intervals of SEPS. Then equal amounts of lysates were analyzed by Western blot analysis using antibody against CXCR4. The results shown here are representative of three independent experiments.
표 Cells were incubated with SEPS (0, 10, 50 or 100 ug/mL) for 24 h.
표 SEPS inhibits CXCR4 by reducing the mRNA level and constitutive activation of NF-kB.
표 (A) SEPS suppresses the expression of CXCR4 mRNA. Cells (1 × 106 cells/well) were treated with 100 ug/mL SEPS for the indicated times. Total RNA was isolated and analyzed by an RT-PCR assay. GAPDH was used to show equal loading of total RNA. The results shown are representative of three independent experiments. (B) Graphs represent band intensities of CXCR4 mRNA. Data was expressed as mean ± SD.
표 SEPS inhibits constitutive NF-kB activation in breast cancer cells.
표 SEPS inhibits binding of NF-kB to the CXCR4 promoter.
표 (A) MDA-MB-231 cells were seeded in the top chamber of a Matrigel. After preincubation with or without SEPS (100 ug/mL) for 6 h, the transwell chambers were placed into 24-well plates with basal medium only or basal medium containing 100 ng/mL CXCL12. After incubation, the cell invasion assay was done. Randomly chosen fields were photographed under a light microscope at 100× magnification and invaded cells were counted. (B) Columns indicate standard average number of invaded cells, bars SE; ** P < 0.01. (C) Invasion assay was performed using the Roche xCELLigence Real-Time Cell Analyzer (RTCA) DP instrument (Roche Diagnostics GmbH, Germany). We tested MDA-MB-231 cells invasion activity in the matrigel-coated CIM (cellular invasion/migration)-Plate 16 with various indicated concentrations of SEPS.
표 Auraptene and osthole inhibits the cell proliferation on human gastric cancer cell lines (A: Auraptene, B: Osthole), (●: AGS, ○: MKN-45, ▼: SNU-1, △: SNU-16, ■: 293T).
표 Effect of auraptene and osthole on cell morphology (A: Auraptene, B: Osthole).
표 Analysis of % of Sub G1 phase in the SNU-1 cells treated with auraptene and osthole.
표 Western blot analysis of apoptosis-related protein expression in auraptene and osthole-treated SNU-1 cells (A: Auraptene, B: Osthole).
표 Inhibition of mammalian target of rapamycin (mTOR) signaling by auraptene leads to an increase of Akt phosphorylation in SNU-1 cells.
표 The nitrite production was measured by the Griess reaction assay.
표 Inhibition of iNOS and COX-2 expressions by OM in LPS-stimulated RAW 264.7 cells.
표 Cells were pretreated with different concentrations of OM for 2 h and stimulated with LPS (1 ug/mL) for 22 h.
표 (A, B, and C) The cells were pretreated with the indicated concentrations of OM for 2 h and stimulated with LPS (1 ug/mL) for 15 min
$(A) The Cells were pretreated with the indicated concentrations of OM for 2 h and stimulated with LPS (1 ug/mL) for 15 min; cytoplasmic and nuclear extracts were prepared resolved by SDS-PAGE, and electrotransferred to a nitrocellulose membrane. (A) After which western blot analyses using IkBa and anti-p65 antibody were performed. b-actin (cytoplasmic fraction) and Lamin B (nuclear fraction) proteins were used as a loading control. The results shown are representative of three independent experiments. (B) After which western blot analyses using c-Jun and c-Fos antibody were performed. Lamin B proteins were used as a loading control. The results shown are representative of three independent experiments.$ 표 (A) The Cells were pretreated with the indicated concentrations of OM for 2 h and stimulated with LPS (1 ug/mL) for 15 min; cytoplasmic and nuclear extracts were prepared resolved by SDS-PAGE, and electrotransferred to a nitrocellulose membrane. (A) After which western blot analyses using IkBa and anti-p65 antibody were performed. b-actin (cytoplasmic fraction) and Lamin B (nuclear fraction) proteins were used as a loading control. The results shown are representative of three independent experiments. (B) After which western blot analyses using c-Jun and c-Fos antibody were performed. Lamin B proteins were used as a loading control. The results shown are representative of three independent experiments.
표 BALB/c mice (ten per group) were injected intraperitoneally with 8 mg/kg of LPS.
표 OM doses of 500 mg/kg were administered orally through an esophageal catheter at 24 h and 2 h before the LPS was injected.
표 Yield of ethanolic extracts from Psidium guajava, Artemisia capillaris Thumb, Coptis chinensis and Taraxacum platycarpum.
표 Effect of ethanolic extracts from Psidium guajava, Artemisia capillaris Thumb, Coptis chinensis and Taraxacum platycarpum on cell viability in RAW 264.7. Each column represents the mean±standard deviation(n=3).
표 Effect of ethanolic extracts from Psidium guajava, Artemisia capillaris Thumb,Coptis chinensis and Taraxacum platycarpum on NO production in LPS- induced RAW 264.7 cells for 24 hr incubation.
표 Effect of ethanolic extracts from Psidium guajava, Artemisia capillaris Thumb,Coptis chinensis and Taraxacum platycarpum on PGE2 production in LPS- induced RAW 264.7 cells for 24hr incubation.
표 Independent variables and their levels used in the response surface design.
표 Central composite design for the optimization of extraction condition for Artemisia capillaris Thumb and Psidium guajava.
표 Polynomial equations calculated by RSM program for extraction conditions of Psidium guajava.
표 Response surface for yield in Psidium guajava extract at constant value (yield:40-55-70%) as a function of extraction temperature, ethanol ratio and extraction time (A). Response surface for total phenolics in Psidium guajava extract at constant value (total phenolics: 68-70-72 mg/100g) as a function of extraction temperature, ethanol ratio and extraction time (B).
표 Response surface for NO production in Psidium guajava extract at constant value (NO production: 50-57.5-65%) as a function of extraction temperature, ethanol ratio and extraction time (A). Response surface for PGE2 production in Psidium guajava extract at constant value (PGE2 production: 50-55-60%) as a function of extraction temperature, ethanol ratio and extraction time (B).
표 Experimental data on yield, total phenolics, NO production and PGE2 production in Psidium guajava extract under different conditions based on central composite design for response surface analysis.
표 Predicted levels of optimum conditions for the maximized and minimized responses of variables by the ridge analysis of their response surface.
표 The range of optimum extraction condition for maximum response variables by superimposing response surface of NO and PGE2 production in Psidium guajava extract.
표 Superimposed response surface for optimization of NO and PGE2 production of Psidium guajava extract as a function of extraction temperature, ethanol ratio and extraction time.
표 Polynomial equations calculated by RSM program for extraction conditions of Artemisia capillaris Thumb.
표 Response surface for yield in Artemisia capillaris Thumb extract at constant value (yield: 20-25-30%) as a function of extraction temperature, ethanol ratio and extraction time (A). Response surface for total phenolics in Artemisia capillaris Thumb extract at constant value (total phenolics: 60-65-68 mg/100g) as a function of extraction temperature, ethanol ratio and extraction time (B).
표 Predicted levels of optimum conditions for the maximized and minimized responses of variables by the ridge analysis of their response surface.
표 The range of optimum extraction condition for maximum response variables by superimposing response surface of NO and PGE2 production in Psidium guajava extract.
표 Superimposed response surface for optimization of NO and PGE2 production of Artemisia capillaris Thumb extract as a function of extraction temperature, ethanol ratio and extraction.
표 Independent variables and their levels used in the response surface design.
표 Central composite design for the optimization of extraction condition for Citrus hassaku pericarp.
표 Total extract yield of ethanolic extracts from Citrus hassaku pericarp.
표 Total phenolic content of ethanolic extracts from Citrus hassaku pericarp.
표 Effect of ethanolic extracts from Citrus hassaku pericarp on cell viability in RAW 264.7. Each column represents the mean±standard deviation(n=3).
표 Response surface for total extract yield in Citrus hassaku pericarp at constant value (yield: 39.0-45.5-51.7%) as a function of extraction temperature, ethanol ratio and extraction time.
표 Response surface for total phenolics in Citrus hassaku pericarp extract at constant value (total phenolics: 37.2-40.0-43.3 mg/g) as a function of extraction temperature, ethanol ratio and extraction time.
표 Response surface for NO production in Citrus hassaku pericarp extract at constant value (NO production: 63.5-72.2-81.7%) as a function of extraction temperature, ethanol ratio and extraction time.
표 Response surface for PGE2 production in Citrus hassaku pericarp extract at constant value (PGE2 production: 61.9-72.2-81.5%) as a function of extraction temperature, ethanol ratio and extraction time.
표 Polynomial equations calculated by RSM program for extraction conditions of Citrus hassaku pericarp.
표 Experimental data on total extract yield, total phenolics, NO production and PGE2 production in Citrus hassaku pericarp extract under different conditions based on central composite design for response surface analysis.
표 Predicted levels of optimum conditions for the maximized and minimized responses of variables by the ridge analysis of their response surface.
표 The range of optimum extraction condition for maximum response variables by superimposing response surface of NO and PGE2 production in Citrus hassaku pericarp extract.
표 Superimposed response surface for optimization of NO and PGE2 production of Citrus hassaku pericarp extract as a function of extraction temperature, ethanol ratio and extraction time.
표 Simplex design plot by mixture design.
표 Effect of ethanolic extracts from mixture (Citrus hassaku pericarp, Psidium guajava, and Artemisia capillaris Thumb extract) on cell viability in RAW 264.7. Each column represents the mean±standard deviation(n=3).
표 Experimental variables and responses used in the simplex-centroid mixture design.
표 Analysis result of the regression and variance for NO production in LPS-induced RAW 264.7 cells.
표 (A) Triangular-dimensional contours diagrams for the effect of Citrus hassaku pericarp (X1), Psidium guajava (X2), and Artemisia capillaris Thumb (X3) on NO production in LPS-induced RAW 264.7 cells. The contour line show % PGE2 production. The contour line show % NO production. (B) Response su(Bace (3D) showing the effect of difB) ent combinations of Citrus hassaku pericarp, Psidium guajava, Artemisia capillaris Thumb extract on NO production in LPS-induced RAW 264.7 cells.
표 Analysis result of the regression and variance for PGE production in2 LPS-induced RAW 264.7 cells.
표 (A) Triangular-dimensional contours diagrams for the effect of Citrus hassaku pericarp (X1), Psidium guajava (X2), and Artemisia capillaris Thumb (X3) on PGE2 production in LPS-induced RAW 264.7 cells. The contour line show % PGE2 production. (B) Response surface (3D) showing the effect of different combinations of Citrus hassaku pericarp, Psidium guajava, Artemisia capillaris Thumb extract on PGE2 production in LPS-induced RAW 264.7 cells.
표 Optimization mixing proportion and composite desirability for responses of NO production and PGE2 production.
표 UPLC-ESI-MS/MS parameters for slected phenolic compounds in the negative mode.
표 Conversion of Animal Doses to Human Equivalent Doses Based on Body Surface Area*.
표 Composition of uncoated Hassaku-guava tablet.
표 Manufacturing process of uncoated Hassaku-guava tablet.
표 Coating's condition for uncoated Hassaku-guava tablet.
표 Quantitative analytical results of the guava leaves extract.
표 MRM chromatograms of standard mixture at 1 μg/mL for target compounds analyzed by negative ionization.
표 MRM chromatograms of the guava leaves extract at 1 μg/mL for target compounds analyzed by negative ionization.
표 Compressed value of uncoated Hassaku-guava tablet.
표 공정별 시료 채취 사진.
표 Total phenolic content(TPC) of manufacturing processes for Hassaku-guava tablet.
표 Microbial contamination levels of manufacturing processes for Hassaku-guava tablet.
표 시제품 사진. 팔삭·구아바 혼합 정제 (A), 팔삭·구아바 혼합음료 (B).

과제명(ProjectTitle) :	-
연구책임자(Manager) :	-
과제기간(DetailSeriesProject) :	-
총연구비 (DetailSeriesProject) :	-
키워드(keyword) :	-
과제수행기간(LeadAgency) :	-
연구목표(Goal) :	-
연구내용(Abstract) :	-
기대효과(Effect) :	-

내보내기 구분	파일저장 인쇄 메일전송
구성항목	기본정보 상세정보 관리번호, 제목(한글), 저자명(한글), 발행일자, 전자원문, 초록(한글), 초록(영문) 관리번호, 제목(한글), 제목(영문), 저자명(한글), 저자명(영문), 주관연구기관(한글), 주관연구기관(영문), 발행일자, 총페이지수, 주관부처명, 과제시작일, 보고서번호, 과제종료일, 주제분류, 키워드(한글), 전자원문, 키워드(영문), 입수제어번호, 초록(한글), 초록(영문), 목차
저장형식	Text(ASCII format) Excel format
메일정보	받는사람 (필수) @ 보내는사람 (선택) @ 제목 내용 KISTI 검색결과 이메일 서비스
안내	총 건의 자료가 검색되었습니다. 다운받으실 자료의 인덱스를 입력하세요. (1-10,000) 검색결과의 순서대로 최대 10,000건 까지 다운로드가 가능합니다. 데이타가 많을 경우 속도가 느려질 수 있습니다.(최대 2~3분 소요) 다운로드 파일은 UTF-8 형태로 저장됩니다. 파일의 내용이 제대로 보이지 않을실 때는 웹브라우저 상단의 보기 -> 인코딩 -> 자동선택 여부를 확인하십시오. ~ Text(ASCII format) Excel format

연합인증

재래귤 유래 항염증 천연활성물질 발굴 및 제품화에 관한 연구
Investigation and commercialization of anti-inflammatory phytochemicals originated from Jeju traditional Citrus 원문보기