표제지
요약
목차
I. 서론 11
II. 재료 및 방법 12
1. 재료 12
2. 시료의 추출 및 부분정제 12
3. 실험동물 14
4. In vitro 시험 14
5. In vivo 시험 15
5.1. 시료의 단회 투여 mouse에 대한 독성 시험 15
5.2. 시료의 복수 암에 대한 mouse 연명효과 시험 15
5.3. 시료의 육종 암에 대한 생육저해효과 시험 16
5.4. 시료 투여에 의한 mouse의 생화학적 시험 17
5.5. Mouse의 조직 검사 17
6. 시료물질의 정제 20
7. 정제물질의 순도측정 및 구조분석 23
III. 결과 24
1. In vitro에서 Sarcoma 180. 세포에 대한 시료의 세포독성 24
2. 단회처리 시료 량에 따른 mouse의 상태변화 26
3. 복수 암 함유 mouse의 시료 경구투여에 따른 연명효과 29
4. 육종 암 함유 mouse의 시료 경구투여에 따른 암 조직 억제 효과 33
5. 시료 경구투여에 따른 육종 암 조직의 HE 및 TUNEL염색 38
6. 육종 암 함유 mo us e의 시료 경구투여에 따른 생리학적 변화 40
7. 육종 암 함유 mouse의 시료 경구투여에 따른 간 및 신장조직의 변화 44
8. 시료물질의 정제 49
9. 정제물질의 구조분석 및 분자량 측정 53
IV. 고찰 60
V. 결론 63
VI. 참고문헌 65
Abstract 68
감사의 글 70
Table 1. IC50 of HaBC18 against Sarcoma 180 cells in vitro 25
Table 2. Mouse mortality by oral medication of HaBC18 27
Table 3. Organ weight in mouse after oral medication of HaBC18 28
Table 4. ILS of mouse bearing acites 180 cells after oral medication of HaBC18 30
Table 5. Body weight of mice bearing acites 180 during oral medication 32
Table 6. Dimensions of a thigh bearing tumor in mouse during oral medication 34
Table 7. TIR of mouse bearing Sarcoma 180 cells after oral medication 36
Table 8. Body weight of mouse bearing tumor during oral medication 41
Table 9. Biochemical properties in blood of mouse bearing tumor after oral medication 42
Table 10. Weight of organs in mouse bearing tumor after oral medication 43
Table 11. Important ¹H-and 13C-NMR Spectral Data of Compound Ha75F1 in Pyridine-d5 57
Fig.1. Morphology of suspending and adherent Sarcoma 180 cells. Suspending(A) and adherent(B) Sarcoma 180 cells before HaBC18 treatment was observed on light microscope. Suspending(A-1) and... 25
Fig.2. Morphological change at the abdominal region of mice after oral medication. A; Healthy mouse, B; Mouse bearing cancer without medication, C; Mouse bearing cancer with adriamycin(0.004g/kg)... 31
Fig.3. Morphology of a thigh bearing tumor in mouse after oral medication. Sarcoma 180 cells(2.5×107cells) were inoculated in a thigh of mouse, and the mice bearing tumor were selected in 9days. Seven mice... 35
Fig.4. Morphology of tumors in mouse after oral medication. Mice were injected with Sarcoma 180 cells(2.5x107cells/100㎕) in a thigh. Seven mice bearing the tumor(MBT) in a group were selected. The... 37
Fig.5. HE and TUNEL staining of tumor cells in mouse after medication. Tumor tissues were staining with HE & TUNEL. NC; MBT without medication, PC1; MBT+adriamycin(0.004g/kg), PC2;... 39
Fig.6. Liver cells in mouse after medication. None; healthy mice, NC; mice bearing tumor without medication, PC1; mice bearing tumor with adriamycin(0.004g/kg), PC2; mice bearing tumor with... 45
Fig.7. HE staining of liver cells in mouse after medication(×300 magnification). A; Mouse bearing tumor(MBT) only, B; Healthy mouse, C; MBT+adriamycin(0.004g/kg), D; MBT+adriamycin(0.002g/kg), E;... 46
Fig.8. Kidney cells in mouse after medication. None; healthy mice, NC; mice bearing tumor without medication, PC1; mice bearing tumor with adriamycin(0.004g/kg), PC2; mice bearing tumor with... 47
Fig.9. HE staining of kidney cells in mouse after medication(×300 magnification). A; Mouse bearing tumor(MBT) only, B; Healthy mouse, C; MBT+adriamycin(0.004g/kg), D; MBT+adriamycin (0.002g/kg), E;... 48
Fig.10. Pattern of HaBC18 by HPLC. HPLC was performed on GS320 column in solvent system(MeOH/MeCN=80/20). The fractions were detected by UV(254nm) and IR detector. 50
Fig.11. Pattern of purified cytotoxic substance(Ha75F1) from HaBC18 by HPLC. HPLC was performed on GS320 column in solvent system(MeOH/MeCN/H₂O=80/20/42). The fractions were detected by UV(254nm) and IR detector. 51
Fig.12. Purified cytotoxic substance(Ha75F1) from HaBC18 on TLC. Solvent system(MeOH/MeCN/H₂O=80/20/42) was developed on RP-C18 TLC. Developed reagent was used Anisadehyde reagent. 52
Fig.13. ¹H-NMR spectrum of Ha75F1. 54
Fig.14. 13C-NMR spectrum of Ha75F1. 55
Fig.15. HMQC spectrum of Ha75F1. 56
Fig.16. MASS spectrum of Ha75F1. 58
Fig.17. Structure of Ha75F1. EI-MS m/z (rel. int, %) 2219[M+] (100) , Molecular formula : C103CH164NO48. 59
List of Schemes
Scheme 1. Procedure of HaBC18 extract from the stem bark of A.julibrissin. 13
Scheme 2. Purification procedure of HaBC18 by HPLC. 22