제주 식물로부터 화장품 관련 유효성분 규명 및 활성 연구
- Alternative Title
- Chemical Constituents and Cosmetics-Related Activities from Plants in Jeju Island
- Abstract
- 본 연구에서는 제주 자생식물을 이용한 기능성 화장품 소재 개발을 목표로 비목나무 (Lindera erythrocarpa Makino), 곰의말채나무 (Cornus macrophylla Wall), 비짜루국화 (Aster subulatus Michx), 넓패 (Ishige sinicola (Setchell et Gardner) Chihara), 참가죽그물바탕말 (Dictyota coriacea (Holmes) Hwang, Kim, et Lee), 쪽동백나무 (Styrax obassia Siebold & Zucc)를 이용한 생리활성성분 규명에 관한 연구를 하였다. 본 연구에 사용된 건조 시료는 70% 에탄올 수용액을 통해 추출되었고, 얻어진 추출물은 극성별로 용매 분획되었다. 각각의 분획층은 활성 검색 (항산화, 항염, 멜라닌합성억제, 항비만)에서 높은 활성을 보이는 분획층을 선택하여 크로마토그래피를 수행 하였다. 이 과정을 통해 35종의 기지 화합물과 2종의 신규 화합물을 분리․동정 하였다. 분리 물질의 화학구조 동정은 고분해능 질량분석기(HR-FAB MS)와 핵자기공명분광기(NMR) 등 분광기기를 통하여 얻어진 데이터를 분석하여 이루어졌다.
비목나무를 이용한 연구 결과, 신규 화합물 1종을 포함한 14종의 성분을 추출물로부터 분리하였다. 분리된 루시돈 (luidone), 메틸린더론 (methyllinderone), 메틸루시돈 (methyllucidone), 1-(2'-hydroxy-3',4',5',6'-tetra-methylphenyl)-1-methoxy-3 -phenylpropane (신규 화합물), 카나쿠지올 (kanakugiol), 린더론 (linderone) 화합물이 흑색종 세포(B16F10)에서 대조군으로 사용된 알부틴과 비교 하였을 때 높은 멜라닌 합성 저해 효과를 나타내었다. 활성을 나타내는 농도에서 화합물이 세포 독성에 영향을 나타내지 않음이 확인되었다. 또한, 멜라닌 합성 mRNA발현 억제실험에서 TRP-1과 티로시나아제 효소 mRNA발현 역시 농도 의존적으로 억제하는 것을 확인하였다.
곰의말채나무로 부터 6종의 성분을 분리하여 동정하였다. 분리된 화합물 중 에틸갈레이트 (ethyl gallate)와 카테킨 (catechin)이 대조군인 BHA와 비교 하였을 때, 자유 라디칼 소거활성이 뛰어남을 확인하였다.
비짜루국화로 부터 1종의 신규화합물과 6종의 기지 물질을 분리 하였다. 분리된 화합물 중 에틸카페에이트 (ethyl caffeate)와 카페인산 (caffeic acid)이 멜라닌합성저해 실험에서 알부틴과 비교 하였을 때 강하게 멜라닌 합성 저해 효과를 나타내었으며, 활성을 나타나는 농도에서 화합물이 세포 독성에 영향을 나타내지 않음을 확인하였다.
갈조류 패과의 해조식물인 넓패 에탄올 추출물로부터 5종의 성분을 분리 하였고, 분리된 화합물 중 리놀레인산 (linoleic acid)은 적은 농도에서도 강하게 멜라닌 합성 저해 효과를 나타내는 것을 확인 하였다. 그리고 분리된 디플로레토하이드록시카마롤 (di-phlorethohydroxycarmalol)이 항비만 실험 중 알파글루코시아다아제 활성 억제 실험에서 대조군으로 사용된 아카보스 (acarbose)와 비교 하였을 때 높은 효소 활성 저해 효과를 나타냄을 확인하였다.
해양 녹조식물인 참가죽그물바탕말의 에탄올 추출물로부터 3종의 성분을 분리 하였다. 분리된 화합물 중 1,9-디하이드록시크레뉴라이드(1,9-dihydroxy -crenulide)와 롤리오라이드 (loliolide)가 강하게 멜라닌 합성 저해 효과를 나타내는 것을 확인 하였다.
쪽동백나무 에탄올 추출물로 부터 2종의 제고사포닌을 분리하였고, 이들은 항비만 효과 실험에서 지방세포분화 (3T3-L1)를 강하게 억제시켜 지방분화를 강하게 억제 시켰으며, 활성을 나타나는 농도에서 화합물이 세포에 독성을 나타내지 않음이 확인되었다.
기능성 화장품 소재개발이 가능한 제주 자생식물로부터 유효 활성 성분을 분리하였고, 관련 분석 장비를 통해 얻은 자료를 토대로 성분의 구조를 동정하였다. 6종의 육․해상 식물로부터 분리된 성분과 생리 활성 연구 결과는 기능성 화장품 소재를 개발하는데 중요한 자료로 활용 될 것이다. 본 연구결과를 통해 총 10건의 국내외 특허를 출원 하였고, SCI급 논문 4편, SCIE급 논문 3편을 등재하였다. 앞으로 본 연구결과를 토대로 심도 깊은 천연물 화학 연구를 계속 진행한다면 더욱 다양한 결과물을 도출할 수 있을 것이라 사료된다.
In this study, investigation into bioactive substances with Lindera erythrocarpa Makino, Cornus macrophylla Wall, Aster subulatus Michx, Ishige sinicola (Setchell et Gardner) Chihara, Dictyota coriacea (Holmes) Hwang, Kim, et Lee, and Styrax obassia Siebold & Zucc was conducted to develop functional cosmetic materials using Jeju native plants. The dried samples were extracted with 70% aq. ethanol and the crude extracts were subjected to solvent fractions according to polarity. Chromatography was carried out for the fractions which showed high biological activities (antioxidant, anti-inflammatory, melanogenesis inhibition, and anti-obesity). Through this process, 37 phytochemicals including 35 known compounds and two new compounds were isolated and identified. Chemical structures of the isolated compounds were identified with analytical data from spectrometers such as HR FAB MS and NMR.
From the extract of Lindera erythrocarpa Makino, 14 phytochemicals including one new compound were isolated. The isolated lucidone, methyllinderone, methyllucidone, 1-(2'-hydroxy-3',4',5',6'-tetramethylphenyl)-1-methoxy-3-phenylpropane (new compound), kanakugiol, and linderone showed high melanogenesis inhibition activity compared with arbutin, the control group, in B16F10 melanoma cell. At the concentration where the activity was appeared, shows activity, the compounds exhibited no cytotoxicity. Furthermore, TRP-1 and tyrosinase mRNA expression were inhibited dose dependently in the suppression experiment of mRNA expression in melanin synthesis.
Six phytochemicals were isolated from Cornus macrophylla Wall and their structures were identified in the same manner as above mentioned. Among the isolates, ethyl gallate and (+)-catechin had significant free radical scavenging effect.
Seven phytochemicals including one new compound were isolated from Aster subulatus Michx. Among the isolates, ethyl caffeate and caffeic acid showed potent melanogenesis inhibition activity without influencing cell toxicities.
Ishige sinicola (Setchell et Gardner) Chihara, Ishigeaceae in Phaeophyta of seaweeds, inhabit in the coast of Jeju. Five phytochemicals were isolated from Ishige sinicola Chihara ethanol extract. Among the isolates, linoleic acid inhibited melanin synthesis strongly at the lower concentration compared to arbutin. Furthermore, isolated di-phlorethohydroxycarmalol showed high enzyme inhibition activity compared to acarbose, the control group of α-glucosidase inhibition activity experiment for anti-obesity.
Three phytochemicals were isolated from ethanol extract of Dictyota coriacea (Holmes) Hwang, Kim, et Lee. Activity screening was conducted using the isolated compounds to investigate melanogenesis inhibition activity and antioxidant effect. As a result, 1,9-dihydroxycrenulide and loliolide showed strong melanogenesis inhibition activity compared to arbutin.
Two Jegosaponins were isolated from ethanol extract of Styrax obassia Siebold & Zucc. For the study of anti-obesity screening, they inhibited on reducing lipid accumulation in 3T3-L1 preadipocytes and formation of cellular lipid contents. The compounds did not exhibit cell toxicities by MTT and LDH assay.
In this study, we have shown that the natural products isolated from Jeju terrestrial and marine plants prossed various biological activities. These results could be utilized to the industrial applications such as functional cosmetic additives in the future.
- Issued Date
- 2011
- Awarded Date
- 2012. 2
- Type
- Dissertation
- Alternative Author(s)
- Ko, Ryeo Kyeong
- Affiliation
- 제주대학교
- Department
- 대학원 화학과
- Advisor
- 이남호
- Table Of Contents
- LIST OF CONTENTS i
LIST OF PHOTOGRAPHS ⅹ
LIST OF SCHEMES ⅹⅰ
LIST OF TABLES ⅹⅱ
LIST OF FIGURES ⅹⅲ
LIST OF ABBREVIATIONS ⅹⅹ
ABSTRACT (In English) ⅹⅹⅳ
초록 (국문) ⅹⅹⅶ
Ⅰ. INTRODUCTION 1
1. Natural product and cosmetics 1
1-1. Need for cosmeceuticals from nature 1
1-2. Botanical extract and cosmetic uses 2
1-3. Characteristics of the phytochemical component used for the cosmetics 5
2. Trend of development of functional cosmetics ingredients 8
2-1. The natural product and functional cosmetic 8
2-2. The overseas technology trend 9
2-3. The domestic technology trend 9
3. Antioxidants 14
3-1. Water-soluble antioxidants 16
3-2. Lipid-soluble antioxidants 18
4. Photoaging agents 20
5. Depigmentation agents 24
5-1. Screening tests for depigmentation agents 25
6. Fat storage and slimming agents 28
7. Research objectives 29
Ⅱ. REAGENT AND INSTRUMENTS 30
1. Chemical Reagent 30
1-1. Column packing material 30
1-2. LC column 30
1-3. TLC (Thin Layer Chromatography) plate 30
1-4. TLC (Thin Layer Chromatography) visualizing reagent 31
1-5. Solvent 31
1-6. HPLC additives for analysis solvent and the preprocessing supplies 31
1-7. Anti-oxidant assay 31
1-8. Anti-inflammatory assay 32
1-9. Melanogenesis inhibition assay 32
1-10. Anti-obesity assay 32
2. Instrument 33
Ⅲ. RESEARCH 1 : Lindera erythrocarpa Makino 34
1. General Plants Information 34
2. Experimental Methods 37
2-1. Plant material 37
2-2. Extraction and solvent fractionation 37
2-2-1. Extraction from the leaves 37
2-2-2. Extraction from the stem barks 37
2-3. Isolation and purification 38
2-3-1. Isolation produce of methylene chloride fraction from the leaves (LM) 38
2-3-2. Isolation produce of ethyl acetate fraction from the leaves (LE) 38
2-3-3. Isolation produce of methylene chloride fraction from the stem barks (SM) 39
3. Qualitative and Quantitative Determination by LC Analysis 42
3-1. HPLC analysis 42
3-1-1. Standard solutions for quantitative analysis 42
3-1-2. Sample preparation for analysis of quantitative and qualitative 42
3-1-3. Determination of HPLC-PDA and ESLD analysis 42
3-1-4. Calibration 44
4. Biological Activity Test 45
4-1. Anti-oxidant assay 45
4-1-1. Preparation of sample 45
4-1-2. DPPH radical scavenging activity assay 45
4-1-3. Superoxide radical scavenging activity assay 46
4-1-4. Xantine oxidase inhibition activity assay 47
4-2. Anti-inflammatory assay 48
4-2-1. Cell culture 48
4-2-2. Cell viability assay by MTT 48
4-2-3. Measurement of NO production 48
4-3. Melanogenesis inhibition activity assay 50
4-3-1. Cell culture 50
4-3-2. Cell viability assay by MTT 50
4-3-3. Measurement of melanin contents 50
4-3-4. Measurement of cell-extracted tyrosinase from cell inhibition activity 51
4-3-5. mRNA preparation and polymerase chain reaction (PCR) 51
4-4. Anti-obesity assay 53
4-4-1. Measurement of yeast α-glucosidase inhibitory activity 53
4-4-2. Cell culture and preadipocyte differentiation 53
4-4-3. Cell viability assay by MTT 54
4-4-4. Cytotoxicity assay (LDH assay) 54
4-4-5. Oil-Red O staining 54
4-5. Statistical analysis 55
5. Results 56
5-1. The structures of the compounds isolated from of L. erythrocarpa 56
5-1-1. Compound 1 56
5-1-2. Compound 2 59
5-1-3. Compound 3 62
5-1-4. Compound 4 65
5-1-5. Compound 5 68
5-1-6. Compound 6 71
5-1-7. Compound 7 74
5-1-8. Compound 8 77
5-1-9. Compound 9 80
5-1-10. Compound 10 83
5-1-11. Compound 11 86
5-1-12. Compound 12 89
5-1-13. Compound 13 92
5-1-14. Compound 14 99
5-2. Biological activities 102
5-2-1. Antioxidant activity 102
5-2-1-1. Free radical scavenging activity of the solvent fractions 102
5-2-1-2. Free radical scavenging activity of the isolated compounds 105
5-2-2. Anti-inflammatory activity 107
5-2-2-1. Effect on cell viability and LPS-induced NO production 107
5-2-3. Melanogenesis inhibition activity 110
5-2-3-1. Cell viability in B16F10 melanoma cell 110
5-2-3-2. Effect on melanogenesis in B16F10 cells 113
5-2-3-3. Morphological observation of B16F10 melanoma cells 116
5-2-3-4. Inhibition effect on melanin synthetic abilities on cell-extracted tyrosinase activity 117
5-2-3-5. Inhibition effect on the related mRNA expression 120
5-2-4. Anti-obesity activity 121
5-2-4-1. Inhibition effect on α-glucosidase 121
5-2-4-2. Cell viability in mouse 3T3-L1 preadipocytes 123
5-2-4-3. Effects on reducing lipid accumulation in mouse 3T3-L1 preadipocytes differentiated adipocytes 126
6. Discussion 130
Ⅳ. RESEARCH 2 : Cornus macrophylla Wall 133
1. General Plants Information 133
2. Experimental Methods 135
2-1. Plant material 135
2-2. Solvent fraction of the leaves 135
2-3. Isolation and purification 135
2-3-1. Isolation produce of ethyl acetate fraction from the leaves (CE) 135
3. Results 137
3-1. The structures of the compounds isolated from of C. macrophylla 137
3-1-1. Compound 1 137
3-1-2. Compound 2 140
3-1-3. Compound 3 143
3-1-4. Compound 4 146
3-1-5. Compound 5 149
3-1-6. Compound 6 150
3-2. Biological activities 151
3-2-1. Antioxidant activity 151
3-2-1-1. Free radical scavenging activity of the crude 70% EtOH extract and the isolated compounds 151
3-2-1-2. Free radical scavenging activity of the isolated compounds 152
4. Discussion 153
Ⅴ. RESEARCH 3 : Aster subulatus Michx 154
1. General Plants Information 154
2. Experimental Methods 156
2-1. Plant material 156
2-2. Extraction and solvent fractionation 156
2-2-1. Extraction from the whole plant 156
2-3. Isolation and purification 156
2-3-1. Isolation produce of methylene chloride fraction (AM) 156
2-3-2. Isolation produce of ethyl acetate fraction (AE) 157
3. Results 159
3-1. The structures of the compounds isolated from of A. subulatus 159
3-1-1. Compound 1 159
3-1-2. Compound 2 160
3-1-3. Compound 3 162
3-1-4. Compound 4 165
3-1-5. Compound 5 172
3-1-6. Compound 6 175
3-1-7. Compound 7 178
3-2. Biological activities 181
3-2-1. Antioxidant activity 181
3-2-1-1. Free radical scavenging activity of the solvent fractions 181
3-2-1-2. Free radical scavenging activity of the isolated compounds 183
3-2-2. Melanogenesis inhibition activity 184
3-2-2-1. Cell viability in B16F10 melanoma cell 184
3-2-2-2. Effect on melanogenesis in B16F10 cells 187
3-2-3. Anti-obesity activity 190
3-2-3-1. Inhibition effect on α-glucosidase 190
3-2-3-2. Cell viability in mouse 3T3-L1 preadipocytes 193
3-2-3-3. Effects on reducing lipid accumulation in mouse 3T3-L1 preadipocytes differentiated adipocytes 194
4. Discussion 195
Ⅵ. RESEARCH 4 : Ishige sinicola (Setchell et Gardner) Chihara 197
1. General Plants Information 197
2. Experimental Methods 199
2-1. Plant material 199
2-2. Solvent fraction from the whole sea plant 199
2-3. Isolation and purification 199
2-3-1. Isolation produce of methylene chloride fraction (IM) 199
2-3-2. Isolation produce of ethyl acetate fraction (IE) 200
3. Results 202
3-1. The structures of the compounds isolated from of I. sinicola 202
3-1-1. Compound 1 202
3-1-2. Compound 2 206
3-1-3. Compound 3 209
3-1-4. Compound 4 212
3-1-5. Compound 5 216
3-2. Biological activities 219
3-2-1. Melanogenesis inhibition activity 219
3-2-2-1. Cell viability in B16F10 melanoma cell 219
3-2-3-2. Effect on melanogenesis in B16F10 cells 222
3-2-3. Anti-obesity activity 224
3-2-3-1. Inhibition effect on α-glucosidase 224
3-2-3-2. Cell viability effect in mouse 3T3-L1 preadipocytes 228
3-2-3-3. Effects on reducing lipid accumulation in mouse 3T3-L1 preadipocytes differentiated adipocytes 231
4. Discussion 234
Ⅶ. RESEARCH 5 : Dictyota Coriacea (Holmes) Hwang, Kim, et Lee 236
1. General Plants Information 236
2. Experimental Methods 238
2-1. Plant material 238
2-2. Solvent fraction from the whole sea plant 238
2-3. Isolation and purification 238
2-3-1. Isolation produce of methylene chloride fraction (DM) 238
3. Results 241
3-1. The structures of the compounds isolated from of D. coriacea 241
3-1-1. Compound 1 241
3-1-2. Compound 2 244
3-1-3. Compound 3 247
3-2. Biological activities 250
3-2-1. Antioxidant activity 250
3-2-1-1. Free radical scavenging activity of the solvent fractions 250
3-2-2. Anti-inflammation activity 252
3-2-2-1. Effect on cell viability and LPS-induced NO production 252
3-2-3. Melanogenesis inhibition activity 254
3-2-3-1. Cell viability in B16F10 melanoma cell 254
3-2-3-2. Effect on melanogenesis in B16F10 cells 257
4. Discussion 260
Ⅷ. RESEARCH 6 : Styrax obassia Siebold & Zucc 262
1. General Plants Information 262
2. Experimental Methods 264
2-1. Plant material 264
2-2. Solvent fraction of the leaves 264
2-3. Isolation and purification 264
2-3-1. Isolation produce of n-butanol fraction (SB) 264
3. Results 268
3-1. The structures of the compounds isolated from of S. obassia 268
3-1-1. Compound 1 268
3-1-2. Compound 2 273
3-2. Biological activities 278
3-2-1. Antioxidant activity 278
3-2-1-1. Free radical scavenging activity of the solvent fractions 278
3-2-2. Melanogenesis inhibition activity 280
3-2-2-1. Cell viability in B16F10 melanoma cell 280
3-2-3-2. Effect on melanogenesis in B16F10 cells 282
3-2-3. Anti-obesity activity 284
3-2-3-1. Inhibition effect on α-glucosidase 284
3-2-3-2. Cell viability in mouse 3T3-L1 preadipocytes 287
3-2-3-3. Effects on reducing lipid accumulation in mouse 3T3-L1 preadipocytes differentiated adipocytes 290
4. Discussion 293
Ⅸ. Overall Conclusion 295
Ⅹ. References 299
Appendix 309
Acknowledgments (감사의 글) 326
- Degree
- Doctor
- Publisher
- 제주대학교 대학원
- Citation
- 고려경. (2011). 제주 식물로부터 화장품 관련 유효성분 규명 및 활성 연구
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