Synthesis and biological activity of phenolic acid conjugated chitosan oligosaccharides
- Alternative Title
- 페놀릭산이 컨쥬게이션된 키토산올리고당의 합성 및 생리 활성
- Abstract
- Phenolic acids are natural hydrophilic antioxidants, which occur ubiquitously in fruits, vegetables, spices, and aromatic herbs. They are of particular interest because of their potential biological properties, such as antioxidant, chelating, free radical scavenging, anti-inflammatory, anti-allergic, anti-microbial, anti-viral, anti-carcinogenic, and UV protection properties. However, due to low solubility of phenolic acids in aqua phase medium, their applications are restricted in the food processing and pharmaceutical industrials.
In this study, eight different substituting groups were introduced on N-2 position of chitosan oligosaccharides (COSs) to potentially improve their functional properties, and chemical structures of the modified derivatives were fully characterized by FT-NMR and UV-Visible spectrophotometer analysis.
In present study, antioxidant activities, apoptosis and cell cycle arrest induced in mouse melanoma cells were investigated. 3,4-dihydroxyl phenolic acid conjugated COSs exhibited the strongest antioxidant activities for reactive oxygen species (ROS), reactive nitrogen species (RNS) and DPPH radicals.
Anti-cancer activity of phenolic acid conjugated COSs was investigated in B16-F10 mouse melanoma cell. PTA-COS showed more effective anti-cancer activity compared to the other samples. In addition, cells treated with PTA-COS showed apoptosis and cell cycle arrest.
- Issued Date
- 2011
- Awarded Date
- 2011. 2
- Type
- Dissertation
- Publisher
- 부경대학교
- Alternative Author(s)
- 엄태길
- Affiliation
- 부경대학교 대학원
- Department
- 대학원 화학과
- Advisor
- 김세권
- Table Of Contents
- Chapter 1 Introduction of chitosan and COSs
1. Background 2
2. The manufacture of COSs 3
3. Biological activities of COSs 7
3.1. Antimicrobial activity 7
3.2. Anti-oxidative effects 9
3.3. Anti-tumor effects 10
4. Research objectives 12
5. Refercences 13
Chapter 2 Preparation and antioxidant effect of phenolic acid conjugated Chitosan oligosaccharides
1. Introduction 22
2. Materials and Methods 26
2.1. Materials 26
2.2. Synthesis of PA-conj-COSs 26
2.3. Instrument analysis 27
2.4. Determination of phenolic acid contents 27
2.5. Reducing power 28
2.6. DPPH radical scavenging assay 28
2.7. Hydroxyl radical scavenging assay 29
2.8. Nitroxide Radical scavenging assay 29
2.9. Intracellular ROS and RNS formation 30
2.10. Statics analysis 30
3. Results 33
3.1. Synthesis and characterization of PA-conj-COSs 33
3.2. Determination of total phenolic content 34
3.3. Reducing power 47
3.3. DPPH radical scavenging activity 49
3.4. Hydroxyl radical scavenging activity 49
3.5. Nitroxide radical scavenging activity 50
3.6. Determination of intracellular ROS and RNS 55
4. Discussion 61
5. References 64
Chapter 3 Protocatechuic acid conjugated COS induce apoptosis and cell cycle arrest in B16-F10 mouse melanoma cells
1. Introduction 73
1.1. Significance of programmed cell death 73
1.2. Extrinsic apoptosis pathways 77
1.3. Instrinic apoptosis pathway 77
1.4. p53 regulated apoptosis 80
1.5. NF-kB regulated cell survival 81
1.6. Cell cycle regulation 82
1.7. Melanoma cancer 87
2. Materials and Methods 89
2.1. Materials 89
2.2. Cell culture 89
2.3. Cytotoxic assessment using WST-1 assay 89
2.4. UV-Vis spectroscopy of enzyme mediated oxidation of PTA-COSs 90
2.5. UV-Vis spectroscopy of Tyrosinase mediated O-quinone formation of PTA-COSs 90
2.6. Cellular ROS Formation 91
2.7. Cellular GSH measurement 91
2.8. Morphological analysis and Hoechst staining 92
2.9. DNA fragmentation assay 92
2.10. Arcridine orange/Ethidium bromide staining 93
2.11. Mitochondria membrane potential (ΔΨm) 93
2.12. Flow cytometry analysis 94
2.13. Western Blot analysis 94
2.14. Statics analysis 95
3. Results 97
3.1. Cytotoxic activity of PA-conj-COSs 97
3.2. Cell proliferation inhibition of PCA-COS 102
3.3. UV-Vis spectroscopy of oxidase mediated oxidation of PTA-COS 102
3.4. Tyrosinase mediated o-quinone formation 107
3.5. Intracellular ROS formation 107
3.6. Intra cellular GSH measurement 108
3.7. Apoptosis determination 108
3.8. Reduction of mitochondrial membrane potential (ΔΨm) in B16-F10 cells treated with PTA-COS 114
3.9. FACS analysis 118
3.10. Effects of PTA-COS on apoptosis related protein expression 118
3.11. PTA-COS induction of p53 protein expression 120
3.12. PTA-COS induces apoptosis via induction of NF-kB protein expression 124
3.13. Effect of PTA-COS on the expression levels of cell cycle regulators 124
3.14. Effect of PTA-COS on the expression levels of MITF signaling 130
3.15. PTA-COS induced Akt and GSK inhibition 131
4. Discussion 136
5. References 144
Chapter 4 Conclusion 159
Abstract in Korean 165
Acknowledgements 167
- Degree
- Doctor
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