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系統識別號 U0007-2807201116255200
論文名稱(中文) 梅花鹿鹿茸改善骨質疏鬆活性研究
論文名稱(英文) Antiosteoporotic Activity of Velvet Antler of Cervus nippon
校院名稱 臺北醫學大學
系所名稱(中) 生藥學研究所
系所名稱(英) Graduate Institute of Pharmacognosy
學年度 99
學期 2
出版年 100
研究生(中文) 宋俊賢
研究生(英文) Chun-Hsien, Sung
學號 M303098014
學位類別 碩士
語文別 中文
口試日期 2011-07-15
論文頁數 76頁
口試委員 指導教授-王靜瓊
委員-林山陽
委員-陳適卿
指導教授-曾頌惠
中文關鍵字 鹿茸  骨質疏鬆  更年期  小鼠骨母細胞      類胰島素生長因子  微型電腦斷層掃描 
英文關鍵字 velvet antler  osteoporosis  menopause  MC3T3-E1  calcium  iron  IGF-1  Micro-CT 
學科別分類
中文摘要 骨質疏鬆症好發於停經後的婦女,主因為女性進入更年期後,卵巢功能降低導致雌激素及黃體素等女性荷爾蒙體內濃度下降,而間接導致骨質的流失。中醫認為更年期和骨質疏鬆與「腎氣」有關;而鹿茸入肝、腎二經,具有壯腎陽,補氣血,益精髓,強筋骨之功效。鹿茸各區段隨著骨化程度之不同,成份及活性都不同。因此本研究以將探討梅花鹿茸之各區段改善骨質疏鬆與成分之關係,找出具預防骨質疏鬆之鹿茸最佳化品質條件。
本研究先以鹿茸乙醇萃取物對MC3T3-E1小鼠骨母細胞進行體外試驗,評估鹿茸萃取物對於骨質重塑之影響。結果顯示:於100 μg/ml以上之鹿茸上段具有最佳刺激MC3T3-E1細胞增生,較控制組增加50%,據Alizarin Red S染色結果顯示25 μg/ml以上,鹿茸上段亦可刺激MC3T3-E1細胞礦化效果,而中段、基部活性依序遞減。繼而分析鹿茸之上段、中段及基部化學分析,結果顯示:蛋白質及總胺基酸含量以上段最多,中段次之,基部最少(蛋白質522、456及391 mg/g;總胺基酸478、437及359 mg/g)。在礦物質方面,鐵是呈現由上段往基部遞減,其含量分別為185、114及92 μg/g,鈣質則由上段往基部遞增,其含量分別為133、164及182 mg/g。其中胰島素樣生長因子(IGF-1)、睪固酮(testosterone)及雌二醇(estradiol)含量亦由上段至基部遞減。
根據衛生署「健康食品之改善骨質疏鬆評估方法」,將卵巢摘除大鼠連續二十週餵服各區段鹿茸(0.1 g/kg/day)。餵服上段之大鼠血中鹼性磷酸酶活性較控制組低,並經Micro-CT分析大鼠脛骨,餵服鹿茸上段、中段之病鼠的骨小樑之骨質比例(BV/TV, Percent bone volume)及單位數量(Tb.N, trabecular number)皆較控制組顯著性增加,而骨小樑分離度(Tb.sp, trabecular separation)較控制組顯著性下降。經骨強度測試儀測試,餵服各區段鹿茸皆使卵巢摘除大鼠的腰椎之強度均優於控制組,而餵服鹿茸上段及中段之病鼠股骨強度則較控制組高。綜合結果,各區段鹿茸均具有改善骨質疏鬆之活性與成分,且以鹿茸上段及中段之活性較佳,配合化學成分分析結果,發現鈣質含量較高之鹿茸骨化程度較高而活性較差,鐵、胺基酸、蛋白質、性激素成分含量較高之鹿茸活性較佳。
英文摘要 Osteoporosis is a disease characterized by low bone mass affecting postmenopausal women. The function of ovary in postmenopausal women is impaired, resulting in decreased estrogen and progesterone level and the bone mass loss. Velvet antler is one of the famous traditional Chinese medicines and has been used in bone disease for a long time. Mineralization is different in degree in different part of velvet antler. With the difference of mineralization, the active components and biological effects of velvet antler may also be different. Therefore, anti-osteoporosis of velvet antler was explored and correlated with the active components and mineralization of different segment of antler in order to suggest future guideline for the quality control of velvet antler.
Velvet antler from sika deers (Cervus nippon) in Quemoy was divided into three sections. In vitro study, MC3T3-E1 cells, an osteoblast precursor cell line were used to evaluate the bone remodeling effect of ethanol extract of velvet antler. The results showed that the top part of velvet antler more significantly stimulated cell proliferation (more than 100 μg/ml) and mineralization (more than 50μg/ml) of MC3T3-E1 cells than the base part. Next, we analyzed the chemical composition of each section. The results show that the amount of protein and total amino acid are richer in the top part, and lesser in the basal part (protein: 522 mg/g, 456 mg/g and 391 mg/g; total amino acid: 478 mg/g, 437 mg/g and 359 mg/g). As for the mineral content analysis, ferrous decreased from top to base (185 μg/g, 114 μg/g and 92 μg/g), but the calcium content increased from base to top, parallel to the degree of mineralization (133 mg/g, 164 mg/g and 182 mg/g). Furthermore, the amount of insulin like growth factor-1(IGF-1), testosterone and estradiol decreased from top to base.
In vivo study, we administered different part of velvet antler (0.1g/kg/day) to the ovariectomized rats orally once daily for 20 weeks. The activity of ALP in the ovariectomized rats fed with the top part of velvet antler is lower than the control. The morphological properties of tibia of rats was evaluated by micro-CT and trabecular separation (Tb. Sp), percent bone volume (BV/TV), trabecular number (Tb.N), and the structure model index (SMI) was calculated. BV/TV, Tb. N and Tb. Sp in the tip and medium section-treated rats were both significantly reversed as compared to control. Using the bone strength tester, the strength of and vertebra was significantly greater in all of the velvet antler treated ovariectomized rats as compared to control. The strength of femur in the tip and medium section-treated rats were both significantly reversed as compared to control.
In conclusion, the study exhibited that each part of velvet antler has the active components to improve osteoporosis effectively, but the top and medium parts are most potent. According to the results of chemical analysis, we suggest the velvet antler shoulder contain less calcium, more iron、glycine、estradiol and IGF-1 in order to have the better anti- osteoporosis effect.
論文目次 摘要..............................................1
Abstract..........................................3
第一章、緒論......................................5
一、骨質疏鬆症簡介................................6
(一)骨骼基本構造..................................6
(二)骨質平衡......................................7
(三)骨質疏鬆症之定義與病因........................9
(四)骨質疏鬆症的臨床診斷與治療....................12
(五)傳統醫學對於骨質疏鬆症之病因與治療............15
二、鹿茸文獻回顧..................................17
(一)鹿茸之簡介....................................17
(二)鹿茸之傳統功效 ................................17
(三)鹿茸之現代藥理研究............................20
(四)鹿茸之化學成份................................21
(五)鹿茸各區段成分差異............................24
三、骨質疏鬆之實驗模式簡介........................26
(一)體外試驗:刺激MC3T3-E1細胞增生及分化模式......26
(二)體內試驗:摘除卵巢誘發之骨質疏鬆疾病模式......27
四、研究動機與目的................................29
五、實驗流程......................................30
第二章、實驗材料與方法............................31
一、實驗材料......................................31
(一)鹿茸樣品......................................31
(二)體外試驗:刺激MC3T3-E1細胞增生及分化模式......31
(三)成分分析......................................32
(四)體內試驗:摘除卵巢誘發之骨質疏鬆疾病模式......33
二、實驗方法......................................35
(一)各區段鹿茸樣品之製備..........................35
(二)體外試驗:刺激MC3T3-E1細胞增生及分化模式......35
(三)成分分析......................................40
(四)體內試驗:摘除卵巢誘發之骨質疏鬆疾病模式......41
三、統計方法......................................44
第三章、結果......................................45
一、鹿茸乙醇萃取物促進MC3T3-E1細胞增生與礦化作用..45
二、各區段鹿茸成分差異............................48
三、建立摘除卵巢大鼠骨質疏鬆模式..................50
四、鹿茸改善大鼠摘除卵巢而誘發之骨質疏鬆效果......55
第四章、討論......................................62
第五章、結論......................................68
第六章、參考資料..................................69
致謝..............................................76
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