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보고서 요약서 3
국문 요약문 4
SUMMARY 5
제1장 연구개발과제의 개요 8
1. 연구개발 목적 8
2. 연구개발의 필요성 8
3. 연구개발 범위 17
제2장 국내외 기술 개발 현황 22
1. 국외 기술, 기술동향 22
2. 국내 기술, 산업동향 22
제3장 연구 수행 내용 및 성과 26
제1세부 과제 : 불가사리를 이용한 발효 최적화 조건 확립 및 항산화 소재 개발 26
1. 연구수행방법 26
2. 연구수행 내용 및 결과 54
제2세부 과제 : 불가사리 발효 추출물 유래 항염증 소재 개발 168
1. 연구수행 방법 168
2. 연구수행 내용 및 결과 176
제4장 목표 달성도 및 관련 분야 기여도 233
제5장 연구개발성과의 활용계획 239
제6장 연구 과정에서 수집한 해외 과학기술 정보 241
제7장 연구개발성과의 보안등급 241
제8장 국가과학기술종합정보시스템에 등록한 연구시설ㆍ장비 현황 241
제9장 연구개발과제 수행에 따른 연구실 등의 안전 조치 이행 실적 241
제10장 연구개발과제의 대표적 연구 실적 242
제11장 기타 사항 242
제12장 참고문헌 243
Table 1. International comparison of aging rate 17
Table 2. Change after 3 months of fertilization 22
Table 3/Table 4. Growth activity of fermented starfish with various mycelia 55
Table 4/Table 5. Extraction yields of fermented starfish with various mycelia 57
Table 5/Table 6. DPPH radical scavenging activity (IC50) 59
Table 6/Table 7. Alkyl radical scavenging activity (IC50) 60
Table 7/Table 8. ABTS radical scavenging activity of fermented starfish using Cordyceps militaris mycelia 62
Table 8/Table 9. ABTS radical scavenging activity fermented starfish using Cordyceps japonica mycelia 63
Table 9/Table 10. Anti-oxidant activity fermented starfish using Cordyceps militaris mycelia 65
Table 10/Table 11. Anti-oxidant activity fermented starfish using Cordyceps japonica mycelia 66
Table 11/Table 12. Growth activity of fermented starfish with various mycelia 70
Table 12/Table 13. Extraction yields of fermented starfish with various mycelia 72
Table 13/Table 14. Fermented starfish with various conditions 73
Table 14/Table 15. Fermented starfish with various conditions 73
Table 15/Table 16. Yield and various radical scavenging activities of fermented starfish and starfish (IC50) 76
Table 16/Table 17. Antioxidant activity, ABTS radical scavenging activity of fermented starfish and starfish 78
Table 17/Table 18. various radical scavenging activities using ESR measurement 83
Table 18/Table 19. Antioxidant activity by ABTS radical scavenging activity and FRAP value 84
Table 19/Table 20. Comparison of radical scavenging activity 85
Table 20/Table 21. Various radical scavenging activity of fermented Asterina pectinifera with Cordyceps militaris mycelia (FACM) and it's fractions 110
Table 21/Table 22. Free amino acid content 117
Table 22/Table 23. Combined amino acid content 118
Table 23/Table 24. Body weight of mice 121
Table 24/Table 25. Liver and spleen weight of mice 122
Table 25/Table 26. Measurement of GOT & GPT from mice serum 124
Table 26/Table 27. Measurement of Triglyceride (TG) from mice serum 125
Table 27/Table 28. Measurement of MDA from mice serum 126
Table 28/Table 29. Measurement of MDA contents of liver 133
Table 29/Table 30. Subjects Assignment 150
Table 30/Table 31. Subjects Information 151
Table 31/Table 32. Distribution on Age 152
Table 32/Table 33. Changes of SOD 153
Table 33/Table 34. Changes of Catalase 154
Table 34/Table 35. Changes of GPx 155
Table 35/Table 36. Changes of TBARS 156
Table 36/Table 37. Changes of Total Cholesterol 158
Table 37/Table 38. Changes of Total Triglyceride 159
Table 38/Table 39. Changes of HDL-Cholesterol 160
Table 39/Table 40. Changes of LDL-Cholesterol 161
Table 40/Table 41. Changes of BUN 163
Table 41/Table 42. Changes of Creatinine 164
Table 42/Table 43. Changes of AST 165
Table 43/Table 44. Changes of ALT 165
Table 44/Table 45. Changes of r-GTP 165
Table 45/Table 46. Changes of ALP 165
Table 46/Table 10. Criteria for histopathological score of osteoarthritic lesions 172
Table 47. 동충하초 밀리타리스 균사체로 발효한 불가사리 추출물의 COX 저해 활성 177
Table 48. 동충하초 자포니카 균사체로 발효한 불가사리 추출물의 COX 저해 활성 178
Table 49. COX-1, COX-2 Inhibition activity 185
Table 50. 소량 발효 추출물과 대량 발효 추출물의 NO 생성 억제 확인 189
Table 51. Effects of SFE treatment on the body weights of papain-induced osteoarthriric mice 199
Table 52. Effects of SFE treatment on the proteoglycan contents in articular cartilages of papain-induced osteoarthritic mice 204
Table 53. Effects of SFE treatment on the histophathological osteoarthritic score of the knee joints of papain-induced osteoarthritic mice 208
Table 54. 주정함유량에 따른 수율 및 라디칼 소거활성 215
Table 55. 50% 주정의 추출시간에 따른 수율 및 라디칼 소거활성 215
Table 56. 추출시간에 따른 수율 및 라디칼 소거활성 216
Table 57. 고속액체크로마토그래프 조건 218
Table 58. 이동상 조건 218
Table 59. 발효 5일차의 HPLC 데이터 223
Table 60. 발효 10일차의 HPLC 데이터 224
Table 61. 발효 15일차의 HPLC 데이터 225
Table 62. 발효 20일차의 HPLC 데이터 226
Table 63. 발효 25일차의 HPLC 데이터 227
Table 64. 시험성적서 요약표 228
Table 65. 영양성분 정보 228
Table 66. Manufacturing process 229
Fig. 1. Environmental pollution caused by starfish 9
Fig. 2. Various diseases caused by free radicals 13
Fig. 3. Global Industry Outlook (2014-2019) 15
Fig. 4. Elderly population ratio trend 16
Fig. 5. Population ratio by age group 17
Fig. 6/Fig. 5. Starfish cream (MIZON) 22
Fig. 7/Fig. 6. Sorabee cosmetic 23
Fig. 8/Fig. 7. Growth activity of fermented starfish with various mycelia 56
Fig. 9/Fig. 8. 수매 불가사리(위)와 채취 불가사리(아래)(원문불량) 67
Fig. 10/Fig. 9. Catching of starfish 68
Fig. 11/Fig. 10. Growth activity of fermented starfish with variety mycelia 71
Fig. 12/Fig. 11. Degree of fermentation of the fermentation period 74
Fig. 13/Fig. 12. Scale-up fermentation(원문불량) 80
Fig. 14/Fig. 13. Observation of the exterior (A) and interior (B) of A. pectinifera fermented with C. militaris mycelia (FACM) 81
Fig. 15/Fig. 14. Cell viability of samples on Chang cells. FACM : fermentd Asterina pertinifera with Cordyceps militaris 87
Fig. 16/Fig. 15. Protective effect of samples on t-BHP-induced oxidative damage in Chang cells. FACM : fermentd Asterina pertinifera with Cordyceps militaris(원문불량) 88
Fig. 17/Fig. 16. Changes of intracellular ROS generated by t-BHP treatment in Chang cells. FACM : fermentd Asterina pertinifera with Cordyceps militaris 89
Fig. 18/Fig. 17. Effects of FACM on cell viability in S16F10 cells. S16FlO cells were treated with indicated concentration of samples for 24 h. Cell viability was... 91
Fig. 19/Fig. 18. Anti-proliferation of FACM on B16F10. (A) Control, (B) FACM (0.05 mg/mL, (C) FACM (0.1 mg/mL), (D) FACM (0.2 mg/mL), 200 X 92
Fig. 20/Fig. 19. Cell cycle analysis of FACM on B16F10. (A) Control, (B) FACM (0.05 mg/mL), (C) FACM (0.1 mg/mL), (D) FACM (0.2 mg/mL) 94
Fig. 21/Fig. 20. Mitochondrial membrane potential (MMP) measurement of FACM in B16F10 cells. (A) Control. (B) FACM (0.05 mg/mL) (C) FACM (0.1 mg/mL) (D) FACM (0.2 mg/mL) 95
Fig. 22/Fig. 21. Tyrosinase inhibitory activity of fermented starfish extract 97
Fig. 23/Fig. 22. Effect of fermented starfish extract on production of melanin in 0.1 μM α-MSH-stimulated B16F10 melanoma cells 98
Fig. 24/Fig. 23. Cell viability of fermented starfish extract 100
Fig. 25/Fig. 24. Protective effect of fermented starfish extract. p〈0.05 vs. normal group, p〈0.05 vs. t-BHP group 101
Fig. 26/Fig. 25. Regulation of ROS production of fermented starfish extract. p〈0.05 vs. normal group, p〈0.05 vs. t-BHP group 102
Fig. 27/Fig. 26. Annexin V/PI staining (A) Control, (B) 100uM t-BHP, (C) 100uM t-BHP, FACM (0.1 mg/mL), (D) 100uM t-BHP + FACM (0.05 mg/mL), (E) 100uM... 104
Fig. 28/Fig. 27. Effect of samples on mitochondrial membrane potential in Chang cells. (A) Control, (B) 100uM t-BHP (C) 100uM t-BHP + FACM (0.1 mg/mL),... 105
Fig. 29/Fig. 28. Spectrum of FACM. FACM : fermentd Asterina pertinifera with Cordyceps militaris 107
Fig. 30/Fig. 29. On-line HPLC profile (a) 210 nm, (b) 230nm (c) 280 nm, (d) 734nm 108
Fig. 31/Fig. 30. Semi-prep HPLC profile 109
Fig. 32/Fig. 31. Analysis HPLC 112
Fig. 33/Fig. 32. LC-Mass peak 113
Fig. 34/Fig. 33. HPLC analysis of cordycepin content. (a) Cordycepin standard (b) Cordyceps militaris mycelia (c) Asterina pectinifera (d) Fermented Asterina... 115
Fig. 35/Fig. 34. Scheme of animal experiment 120
Fig. 36/Fig. 35. Liver of mice after sacrifice. (A) normal mice, (B) t-BHP (2 mmol/kg) treated mice, (C) FACM (25 mg/kg), (D) FACM (50 mg/kg) with t-BHP (E)... 128
Fig. 37/Fig. 36. Effects of FACM on SOD activity in t-BHP-induced oxidative stress mice. Normal: normal mice. Damage: t-BHP (2 mmol/kg) treated mice, Sample... 129
Fig. 38/Fig. 37. Effects of FACM on Catalase activity in t-BHP-induced oxidative stress mice. Normal: normal mice. Damage: t-BHP (2 mmol/kg) treated mice,... 130
Fig. 39/Fig. 38. Effects of FACM on total glutathione activity in t-BHP-induced oxidative stress mice. Normal: normal mice. Damage: t-BHP (2 mmol/kg) treated... 131
Fig. 40/Fig. 39. H&E staining (A) Normal: normal mice. (B) Damage: t-BHP (2 mmol/kg) treated mice, (C) Low concentration: t-BHP and low concentration of sample... 134
Fig. 41/Fig. 40. 전문가 자문 140
Fig. 42/Fig. 41. 방문상담 및 모둠토의(원문불량) 141
Fig. 43/Fig. 42. Effect of the FACM on the expression of Bcl-2, Bax in Chang cells 143
Fig. 44/Fig. 43. Effect of the FACM on the expression of various caspases in the apoptotic pathway in Chang cells 144
Fig. 45/Fig. 44. Apoptosis pathway 145
Fig. 46/Fig. 45. Compare the expression of various caspases in the apoptotic pathway 147
Fig. 47/Fig. 46. Cell viabilities of NHDF cells treated C.militaris mycelium fermented starfish extracts 179
Fig. 48/Fig. 47. Cell viabilities of NHDF cells treated P. Japonica fermented starfish extracts 180
Fig. 49/Fig. 48. Effect of C.militaris mycelium fermented starfish extracts on production of IL-6 in RAW264.7 cells 181
Fig. 50/Fig. 49. Effect of P. Japonica fermented starfish extracts on production of IL-6 in RAW264.7 cells 182
Fig. 51/Fig. 50. Effect of C.militaris mycelium fermented starfish extracts on production of TNF-α in RAW264.7 cells 183
Fig. 52/Fig. 51. Effect of P. Japonica fermented starfish extracts on production of TNF-α in RAW264.7 cells 184
Fig. 53/Fig. 52. Effect of starfish extracts on production of nitric oxide in RAW264.7 cells 186
Fig. 54/Fig. 53. Cell viabilities of RAW264.7 cells treated fermented starfish extracts 187
Fig. 55/Fig. 54. Effect of fermented starfish extracts on production of nitric oxide in RAW264.7 cells 188
Fig. 56/Fig. 55. HPLC 분석 결과 191
Fig. 57/Fig. 56. 분취물별 세포 독성 분석 결과 192
Fig. 58/Fig. 57. 분취물별 항염증 분석 결과(NO 활성 억제) 193
Fig. 59/Fig. 58. 분취물별 항염증 분석 결과(IL-6 활성 억제) 195
Fig. 60/Fig. 59. 18분 peak의 항염증 분석 결과(COX-1, COX-2) 196
Fig. 61/Fig. 60. 22분 peak의 COX-1, COX-2 분석 결과 197
Fig. 62/Fig. 61. Effects of SFE on the TNF-α contents in plasma of papain-induced osteoarthriric mice. Each value is expressed as mean±SD. Values with different... 200
Fig. 63/Fig. 62. Effects of SFE on the IL-1β contents in plasma of papain-induced osteoarthriric mice. Each value is expressed as mean±SD. Values with different... 201
Fig. 64/Fig. 63. Effects of SFE on the IL-6 contents in plasma of papain-induced osteoarthriric mice. Each value is expressed as mean±SD. Values with different... 202
Fig. 65/Fig. 64. Safranin O stain intensity of articular cartilage of tibial platues. (A) Stain intensity of red color increased in proportion to proteoglycan content. (B)... 205
Fig. 66/Fig. 65. Microphotograph of articular structure of knee joint. (A) In normal group, articular cartilages are well developed. (B) In papain group, articular... 207
Fig. 67/Fig. 66. ALT after SFE treatment for 28 days in papain-induced osteoarthritic mice. Each value is expressed as mean±SD. Values with different alphabets... 210
Fig. 68/Fig. 67. AST after SFE treatment for 28 days in papain-induced osteoarthritic mice. Each value is expressed as mean±SD. Values with different alphabets... 211
Fig. 69/Fig. 68. BUN after SFE treatment for 28 days in papain-induced osteoarthritic mice. Each value is expressed as mean±SD. Values with different alphabets... 212
Fig. 70/Fig. 69. Creatinine after SFE treatment for 28 days in papain-induced osteoarthritic mice. Each value is expressed as mean±SD. Values with different... 213
Fig. 71/Fig. 70. Manufacturing process steps 229
Fig. 72/Fig. 71. 임상시험용 캡슐 사진 230
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