Title Page
Contents
List of Abbreviations 13
Abstract 16
Chapter 1. Literature Review 18
1.1. Ginger (Zingiber officinale) leaf 18
1.2. Upcycling 21
1.2.1. Sustainable Food System 21
1.2.2. Food by-products 22
1.2.3. Upcycling of agricultural food by-products 24
1.3. Enzyme-Assisted Extraction (EAE) 27
1.4. Response surface method (RSM) 31
1.5. Immunity in RAW264.7 34
Chapter 2. Introduction 36
Chapter 3. Optimization of Enzyme-Assisted Extraction 38
3.1. Materials & Methods 38
3.1.1. Plant materials 38
3.1.2. Chemicals and enzyme 38
3.1.3. Preparation of ginger leaf extracts 39
3.1.4. Extraction yield of sample extracts 41
3.1.5. Water solubility index (WSI) 41
3.1.6. Total phenolic acid content (TPC) 42
3.1.7. Total Flavonoid content (TFC) 42
3.1.8. DPPH radical scavenging activity 43
3.1.9. ABTS radical scavenging activity 43
3.1.10. Experimental design 44
3.1.11. Statistical analysis 46
3.2. Result & Discussion 47
3.2.1. Selecting the enzyme for optimization 47
3.2.2. Model fitting analysis by BBD 49
3.2.3. Effect of EAE factors on extraction yield 53
3.2.4. Effect of EAE factors on TPC 56
3.2.5. Effect of EAE factors on TFC 58
3.2.6. Effect of EAE factors on ABTS radical scavenging activity 60
3.2.7. Effect of EAE factors on DPPH radical scavenging activity 62
3.2.8. Optimization of the EAE conditions and its verification 64
3.2.9. Optimized Enzyme-assisted extract versus control extract 66
Chapter 4. Immunomodulating Activity Effects of ginger leaf in RAW264.7 68
4.1. Materials & Methods 68
4.1.1. Materials 68
4.1.2. Chemicals 68
4.1.3. Cell Culture 69
4.1.4. Cell viability assay 69
4.1.5. Nitric oxide assay 70
4.1.6. Enzyme-linked immunosorbent assay (ELISA) 70
4.1.7. Western blot analysis of immune-related enzymes 71
4.1.8. Statistical analysis 72
4.2. Result & Discussion 73
4.2.1. Effect of ginger leaf EAE on cell viability in RAW264.7 cells. 73
4.2.2. Effect of ginger leaf EAE on nitric oxide (NO) production of RAW264.7 cell 75
4.2.3. Effect of ginger leaf EAE on Cytokine production in RAW264.7 cells. 77
4.2.4. Cytokine-producing immune mediators were upregulated by ginger leaf EAE treatment. 83
4.3. Conclusion 87
Abstract (in Korean) 88
Reference 90
Table 1. Characteristics of enzymes used in this study 30
Table 2. Box-Behnken design independent variable values and corresponding levels 45
Table 3. Antioxidant measurement result of ginger leaf extract according to enzyme type 48
Table 4. Experimental design of extraction parameters for the optimization of TPC, TFC, DPPH and ABTS, Yield of Zinglber officinale Leaf extracts obtained with EAE 51
Table 5. Polynomial regression equation for the total phenolic acid contents, total flavonoid contents, DPPH radical scavenging activity, ABTS radical scavenging activity, extraction yield from ginger leaf extract 52
Table 6. Optimum extraction conditions and validation of predicted and experimental values 65
Table 7. Comparison of antioxidant activity and yield of optimal condition extract of the ginger leaf (EAE) vs. enzyme-treated extract of the ginger leaf (control, C-EAE) 67
Figure 1. ginger (Zingiber officinale) (a) leaf, (b) stem and root. 20
Figure 2. Upcycling of agricultural food by-products. 26
Figure 3. Box-Behnken design model. 33
Figure 4. preparation of ginger leaf extract and procedure of selecting optimal enzyme. 40
Figure 5. Response surface plots of yield (%) of extract methods. 55
Figure 6. Response surface plots of total phenolic acid contents (mg GAE/g) of extract methods. 57
Figure 7. Response surface plots of total flavonoid contents (mg CTE/g) of extract methods. 59
Figure 8. Response surface plots of ABTS radical scavenging activity (% inhibition) of extract methods. 61
Figure 9. Response surface plots of DPPH radical scavenging activity (% inhibition) of extract methods. 63
Figure 10. Effects of ginger leaf extracts on the production of Cell Viability in RAW264.7. 74
Figure 11. Effects of ginger leaf extracts on the production of nitric oxide in RAW264. 76
Figure 12. Effects of ginger leaf extracts on the production of Cytokine TNF-α in RAW264.7. 79
Figure 13. Effects of ginger leaf extracts on the production of Cytokine IL-6 in RAW264.7. 80
Figure 14. Effects of ginger leaf extracts on the production of Cytokine IL-1β in RAW264.7. 81
Figure 15. Effects of ginger leaf extracts on the production of Cytokine PGE2 in RAW264.7. 82
Figure 16. COX-2 protein expressions in RAW264.7 cells by GL samples. 85
Figure 17. iNOS protein expressions in RAW264.7 cells by GL samples. 86