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系統識別號 U0002-2507201210293000
DOI 10.6846/TKU.2012.01082
論文名稱(中文) 幾丁聚醣/磷酸鈣複合材料之製備及作為引導骨再生薄膜之評估
論文名稱(英文) Preparation of chitosan/calcium phosphate composite materials useful for the guided bone regeneration
第三語言論文名稱
校院名稱 淡江大學
系所名稱(中文) 化學工程與材料工程學系博士班
系所名稱(英文) Department of Chemical and Materials Engineering
外國學位學校名稱
外國學位學院名稱
外國學位研究所名稱
學年度 100
學期 2
出版年 101
研究生(中文) 戴宏穎
研究生(英文) Hung-Yin Tai
學號 695400365
學位類別 博士
語言別 繁體中文
第二語言別
口試日期 2012-07-17
論文頁數 178頁
口試委員 指導教授 - 董崇民(tmdon@mail.tku.edu.tw)
委員 - 鄭廖平(lpcheng@mail.tku.edu.tw)
委員 - 林達鎔(djlin@mail.tku.edu.tw)
委員 - 孫一明(cesunym@saturn.yzu.edu.tw)
委員 - 傅鍔(ndmcearl@ndmctsgh.edu.tw)
委員 - 何明樺(mhho@mail.ntust.edu.tw)
委員 - 蔡敏郎(tml@mail.ntou.edu.tw)
關鍵字(中) 幾丁聚醣
磷酸鈣
引導骨再生
關鍵字(英) chitosan
calcium phosphate
guided bone regeneration
asymmetric composite membrane
第三語言關鍵字
學科別分類
中文摘要
本研究利用幾丁聚醣作為引導骨再生膜的主材料,幾丁聚醣具有良好細胞相容性、無毒性、生物可分解性、成膜性以及價格便宜等特性,加上本身具有抑菌性,可防止傷口細菌感染。而為了加速引導骨再生,另外在幾丁聚醣中加入不同結晶型態之磷酸鈣混合成複合材料,藉由磷酸鈣不斷釋放出鈣離子來引導骨性細胞吸附及加速骨組織的再生,同時亦藉著磷酸鈣來改善幾丁聚醣的機械性質。本研究乃利用逆乳化法製備缺鈣之氫氧基磷灰石(CDHA),然後經800°C與1000°C高溫煆燒後,可分別轉換成結晶性較高且同時含有缺鈣氫氧基磷灰石及磷酸三鈣(TCP)的雙相磷酸鈣(BCP)與磷酸三鈣(TCP),將不同的磷酸鈣(CDHA, BCP, TCP)引入幾丁聚醣之中,並以冷凍乾燥法製備成多孔結構薄膜(chitosan-P, CDHA/chitosan-P, BCP/chitosan-P, TCP/chitosan-P),適當的孔洞尺寸與孔隙度,可以讓細胞或組織的增生能力提高,但使其不易侵入薄膜而破壞薄膜結構。實驗發現,骨母細胞較易貼附於含有CDHA結晶相態的CDHA/chitosan-P與BCP/chitosan-P孔洞薄膜上,而骨母細胞在具有TCP相態的BCP/chitosan-P與TCP/chitosan-P孔洞薄膜上則有較強的細胞增生能力,最終骨母細胞分化與礦化時,CDHA/chitosan-P與 BCP/chitosan-P孔洞薄膜中的CDHA結晶相態又可引導骨母細胞分化形成新生骨礦物,總結來說,冷凍乾燥法製備之薄膜具有的多孔結構,可讓牙齦纖維母細胞在薄膜的一側生長且無不良反應,而對另一側骨缺損的部份,又有加強其骨細胞再生的能力。其中又以BCP/chitosan-P因同時擁有CDHA與TCP兩結晶相態,讓BCP/chitosan-P具備與骨母細胞貼附相關的骨親合性,同時還具有加速骨母細胞增生,與增強骨母細胞分化礦化的能力,因此在引導骨再生材料上,BCP/chitosan-P確實擁有其應用發展的空間。將幾丁聚醣溶液直接冷凍乾燥可製備得到單純孔洞型態之幾丁聚醣薄膜,其材料較為柔軟,操作性較不佳。為了這個缺失,本研究在幾丁聚醣孔洞膜的一側加上一層生物可分解聚羥基丁酯(PHB)緻密層,將PHB薄膜電漿接枝聚丙烯酸,並以聚丙烯酸為架橋固定幾丁聚醣,直接將幾丁聚醣冷凍乾燥製備出非對稱型幾丁聚醣/PHB複合薄膜,此種薄膜不論在乾燥或濕潤的狀態下,材料皆具有優異的起始模數,最終,在動物實驗方面,本研究比較不同磷酸鈣(CDHA, BCP與TCP)與幾丁聚醣製備之多孔膜,將材料貼附在大鼠頭蓋骨缺損位置21天後,可以發現BCP/chitosan-P薄膜覆蓋在骨缺損部位後,可以在21天內將骨缺損恢復將近58%,大幅的加速骨修復之速度。本研究所製備之添加BCP顆粒之幾丁聚醣孔洞薄膜確實具有在引導骨組織再生膜的應用發展之空間。
英文摘要
To fulfill the properties of barrier membrane useful for guided bone tissue rgegenration in the treatment of periodontitis, a simple process containing oven-drying and lyophilization was proposed in this study to produce composite membrane from biodegradable chitosan (CS) and fnctional calcium phosphate. Calcium deficient hydroxyapatite (CDHA) having an average particle size of 45 nm was synthesized by reverse emulsion method. It was converted to the respective biphasic calcium phosphate (BCP, 226 nm) and ß-tricalcium phosphate (TCP, 450 nm) by calcination at 800°C and 1000°C, and the BCP consisted of 92% TCP and 8% CDHA. Subsequently, chitosan was mixed with calcium phosphates to prepare CDHA/chitosan, BCP/chitosan, and TCP/chitosan membranes. The lyophilization at -20°C then gave the rest of material an interconnected pore structure with high porosity (91~95%) and suitable pore size (102 to 147 μm) which could reveal the biocompatibility and prevention of migration of gingival fibroblast. Furthermore, the composite membrane promoted the permeability and adhesiveness to bone cells as demonstrated by the in-vitro cell culture of primary osteoblast. The CDHA and BCP particles separately added into chitosan could further increase the cell attachment and differentiation of osteoblast, and then the BCP and TCP particles separately added into chitosan could increase great cell proliferation of osteoblast. This is beneficial for maintaining a secluded space for the bone regeneration as well as to prevent the invasion of other tissues. On the other hand, asymmetric chitosan/PHB composite membrane was produce to promote the wet operation of barrier membrane. The dense PHB membrane grafted poly(acrylic acid) by plasma induced polymerization, then chitosan was immobilized by reacting with poly(acrylic acid) and lyophilized. The asymmetric membrane (chitosan/PHB) with a skin PHB layer had initial modulus value almost 50 times that of the symmetric porous membrane (chitosan-P). Finally, five different samples of a control, chitosan, CDHA/chitosan, BCP/chitosan, and TCP/chitosan porous membrane were assessed as periodontal barrier membranes for the calvarial critical size bone defects in SD rats. Histological and histomorphometric analyses revealed that BCP/chitosan resulted in the most effective bone regeneration compared to other samples with rgenerated bone area of 57% covering the bone defect area. The BCP/chitosan has the potential to be used as the barrier membrane for guided bone regeneration.
第三語言摘要
論文目次
目錄
第一章 緒論	1
第二章 文獻回顧	3
2-1 引導組織再生工程	3
2-2 生醫材料基材特性	7
第三章 實驗藥品與實驗儀器	13
3-1 實驗藥品	13
3-2 實驗儀器	15
第四章 以逆乳化法製備磷酸鈣之製程探討與其特性研究	17
4-1 緒論	17
4-2 實驗流程與方法	20
4-2.1 以逆乳化法合成磷酸鈣	20
4-2.2 磷酸鈣奈米顆粒結構型態與顆粒特性分析	21
4-3 實驗結果與討論	25
4-3.1 以逆乳化法合成磷酸鈣顆粒與不同煆燒溫度下之結晶構造分析	25
4-3.2 不同結晶型態之磷酸鈣的溶解行為與其結晶變化	40
4-3.3 不同結晶型態磷酸鈣對骨母細胞之貼附(cell adhesion)與增生(cell proliferation)之活性影響	43
4-4 結論	47
第五章 磷酸鈣/幾丁聚醣複合薄膜製備與其特性研究	48
5-1 緒論	48
5-2 實驗流程與方法	51
5-2.1 幾丁聚醣的純化及鑑定	51
5-2.2 幾丁聚醣-鈣磷酸鹽複合薄膜之製備	53
5-2.3 磷酸鈣/幾丁聚醣複合薄膜結構與性質分析	54
5-2.4 幾丁聚醣複合薄膜之細胞毒性測試與引導組織再生能力評估	56
5-3 實驗結果與討論	63
5-3.1 幾丁聚醣的性質測定	63
5-3.2 磷酸鈣/幾丁聚醣複合薄膜的製備	66
5-3.3 磷酸鈣/幾丁聚醣複合薄膜複合薄膜結晶構造分析	71
5-3.4 磷酸鈣/幾丁聚醣複合薄膜的酵素水解	73
5-3.5 In vitro磷酸鈣/幾丁聚醣複合薄膜在模擬體液(SBF)中的變化	81
5-3.6 In vitro磷酸鈣/幾丁聚醣複合薄膜的細胞毒性測試	86
5-3.7 磷酸鈣/幾丁聚醣複合薄膜應用於引導骨再生的模擬	88
5-4 結論	99
第六章 非對稱性複合薄膜製備與其特性研究	100
6-1 緒論	100
6-2 實驗流程與方法	103
6-2.1 利用預烘/凍乾法製備非對稱幾丁聚醣薄膜的製備	103
6-2.2 PHB的純化與薄膜製備	104
6-2.3 PHB薄膜的電漿接枝聚合	105
6-2.4 非對稱型複合薄膜結構與性質分析	106
6-2.5 幾丁聚醣複合薄膜之細胞毒性評估	107
6-3 實驗結果與討論	110
6-3.1 非對稱型磷酸鈣/幾丁聚醣複合薄膜的製備	110
6-3.2 非對稱型幾丁聚醣/PHB複合薄膜之製備	117
6-3.3 對稱性與非對稱性幾丁聚醣複合薄膜的機械性質	123
6-3.4 幾丁聚醣複合薄膜之細胞相容性	128
6-4 結論	131
第七章 不同磷酸鈣/幾丁聚醣複合材料之引導骨再生評估	132
7-1 緒論	132
7-1.1 骨組織的組成	132
7-1.2 骨瘉合的過程 	133
7-1.3 骨組織工程	135
7-2 實驗流程與方法	136
7-2.1 幾丁聚醣複合薄膜之阻隔及引導組織再生能力評估	136
7-2.2 動物實驗	140
7-3 實驗結果與討論	142
7-3.1 非對稱型幾丁聚醣複合薄膜對牙齦纖維母細胞之活性與阻隔性	142
7-3.2 非對稱型幾丁聚醣複合薄膜的引導骨細胞再生能力	150
7-3.3 動物實驗	153
7-4 結論	162
第八章 結論	163
參考文獻	165
 
圖目錄
圖2-1 牙周炎示意圖	3
圖2-2 引導組織再生示意圖	4
圖2-3 可吸收性引導組織再生示意圖	5
圖2-4 生醫材料的分類示意圖	8
圖2-5 陶瓷材料的特性圖	8
圖4-1 逆乳化法中鈣與磷離子在微胞中的反應示意圖	20
圖4-2 逆乳化法合成磷酸鈣實驗流程示意圖	21
圖4-3 恆溫振盪示意圖	22
圖4-4 磷酸鈣前處理與種植細胞示意圖	25
圖4-5 將加入4 vol% Span80的cyclohexane混合液分別加入鈣與磷前驅物水溶液以超音波震盪得到的逆乳化微胞(a)Ca(NO3)2、(b)(NH4)2HPO4與(c)將兩乳液混合再次超音波震盪之微胞,以及利用不同Span80濃度所合成之乾燥磷酸鈣顆粒(d)之粒徑分析圖	27
圖4-6為不同起始鈣磷比(Ca/P)i之逆乳化合成產物與其經過高溫煆燒產物之XRD圖譜	30
圖4-7 以逆乳化法合成磷酸鈣與再經不同高溫煆燒之產物結晶構造和不同起始鈣磷比(Ca/P)i之關係圖,結晶構造是以XRD圖譜經DIFFRACplus EVA軟體判別。	31
圖4-8 不同起始鈣磷比(Ca/P)i與其逆乳化反應生成之產物鈣磷比(Ca/P)p關係圖	32
圖4-9 以逆乳化法合成磷酸鈣在不同反應時間下所得產物之TEM圖與其產率關係圖	34
圖4-10 以逆乳化法合成出之CDHA及在800oC與1000oC煆燒所獲得之BCP與TCP產物的TEM圖與其粒徑分析圖	35
圖4-11 以逆乳化法合成之CDHA於800oC下煆燒不同時間所得產物之TEM比較圖	36
圖4-12 以逆乳化法合成出之CDHA及在800oC與1000oC煆燒所獲得之BCP與TCP的FTIR圖	38
圖4-13 以逆乳化法合成出之CDHA及在800oC與1000oC煆燒所獲得之BCP與TCP之XRD圖	39
圖4-14 以逆乳化法合成出之CDHA之 TG-DSC分析	40
圖4-15 以逆乳化法合成出之CDHA及在800oC與1000oC煆燒所獲得之BCP與TCP於去離子水中的鈣離子溶解曲線圖	42
圖4-16 以逆乳化法合成出之CDHA及在800oC與1000oC煆燒所獲得之BCP與TCP於去離子水浸泡不同天數後XRD變化圖	43
圖4-17 骨母細胞於CDHA, BCP and TCP粉末上培養一天後的DAPI螢光分析(a), (b)及(c),以及(d)細胞檢量線和(e)培養1-4天之細胞生長曲線圖	46
圖5-1幾丁聚醣的結構式	48
圖5-2 磷酸鈣/幾丁聚醣複合薄膜之製備流程圖	53
圖5-3 (a)不同濃度N-乙醯基葡萄糖胺溶液的一次微分紫外線吸收光譜圖;(b)不同濃度N-乙醯基葡萄糖胺溶液在203 nm波長的一次微分值對濃度作圖之檢量線	64
圖5-4 幾丁聚醣溶液的還原黏度( )與固有黏度( )對濃度作圖	66
圖5-5 以乾式製程製備之chitosan-D, CDHA/chitosan-D, BCP/chitosan-D and TCP/chitosan-D的上表面SEM圖	67
圖5-6 以冷凍乾燥法製備之chitosan-P, CDHA/chitosan-P, BCP/chitosan-P and TCP/chitosan-P的(a)上表面與(b)下表面SEM圖	69
圖5-7 以冷凍乾燥法製備之chitosan-P, CDHA/chitosan-P, BCP/chitosan-P and TCP/chitosan-P孔洞薄膜浸入林格氏液後的吸水曲線圖	70
圖5-8 (a)乾式製程製備chitosan-D、CDHA/chitosan-D、BCP/chitosan-D及TCP/chitosan-D薄膜的XRD圖,(b)冷凍乾燥製程製備chitosan-P、CDHA/chitosan-P、BCP/chitosan-P及TCP/chitosan-P薄膜的XRD圖。	72
圖5-9 比較低分子量幾丁聚醣孔洞膜(LMW chitosan-P, MW=170 kDa)在不同酵素液之中酵素(pH=7; T=37oC)水解情況。■2 mg/mL amylase and 0.5 mg/mL lysozyme;▲2 mg/mL amylase;●0.5 mg/mL lysozyme	76
圖5-10 高分子量幾丁聚醣孔洞膜(HMW chitosan-P, MW=509 kDa))與低分子量幾丁聚醣孔洞膜(LMW chitosan-P, MW=170 kDa)於酵素水解(pH=7; T=37oC)上的差異,酵素液為2 mg/mL amylase and 0.5 mg/mL lysozyme	77
圖5-11 (a)磷酸鈣/幾丁聚醣緻密薄膜與(b)磷酸鈣/幾丁聚醣孔洞薄膜的酵素水解(pH=7; T=37oC)重量損失圖,以原始磷酸鈣/幾丁聚醣複合薄膜總重作為基準,而2 mg/mL amylase and 0.5 mg/mL lysozyme為酵素液。	79
圖5-12 (a)磷酸鈣/幾丁聚醣緻密薄膜與(b)磷酸鈣/幾丁聚醣孔洞薄膜的酵素水解(pH=7; T=37oC),經過14天後表面SEM觀察圖,以2 mg/mL amylase and 0.5 mg/mL lysozyme為酵素液	80
圖5-13 將不同磷酸鈣/幾丁聚醣孔洞薄膜的水解損失扣除磷酸鈣溶解部份(圖4-15)後得到的水解損失圖,以原始複合薄膜中的幾丁聚醣重量作為基準。以2 mg/mL amylase and 0.5 mg/mL lysozyme為酵素液,酵素水解環境控制於pH=7; T=37oC。	81
圖5-14 磷酸鈣/幾丁聚醣複合薄膜(a)浸入去離子水時,去離子水中鈣離子濃度變化圖與 (b)浸入模擬體液時,模擬體液中鈣離子濃度變化圖	83
圖5-15 磷酸鈣/幾丁聚醣複合薄膜於浸泡模擬體液後的XRD圖譜	84
圖5-16 磷酸鈣/幾丁聚醣複合薄膜浸泡過模擬體液後的表面SEM圖(a)緻密薄膜與(b)孔洞薄膜	85
圖5-17 L929種植於(a)磷酸鈣/幾丁聚醣緻密薄膜與(b)磷酸鈣/幾丁聚醣孔洞薄膜的細胞生長曲線	87
圖5-18 人類牙齦纖維母HGF種植於(a)磷酸鈣/幾丁聚醣緻密薄膜與(b)磷酸鈣/幾丁聚醣孔洞薄膜的細胞生長曲線	89
圖5-19 骨母細胞OB種植於(a) 磷酸鈣/幾丁聚醣緻密薄膜與(b) 磷酸鈣/幾丁聚醣孔洞薄膜初期1-6小時之細胞貼附曲線	91
圖5-20 骨母細胞OB種植於磷酸鈣/幾丁聚醣孔洞薄膜4和6小時後的DIPA螢光圖	92
圖5-21 骨母細胞OB種植於(a)磷酸鈣/幾丁聚醣緻密薄膜與(b)磷酸鈣/幾丁聚醣孔洞薄膜初期1-4天之細胞增生曲線	94
圖5-22 骨母細胞OB種植於磷酸鈣/幾丁聚醣孔洞薄膜2天後,SEM觀察圖	95
圖5-23 (a)磷酸鈣/幾丁聚醣孔洞薄膜上的骨母細胞在不同天數的ALP活性圖與(b)骨母細胞礦物化之鈣離子含量	97
圖5-24 骨母細胞OB於磷酸鈣/幾丁聚醣孔洞薄膜上誘導分化21天後,薄膜SEM觀察圖,白色箭頭所指位置即為骨小結沉積部份	98
圖5-25 骨母細胞OB於磷酸鈣/幾丁聚醣孔洞膜上誘導分化21天後,薄膜XRD觀察圖	98
圖6-1 不同結構型態之薄膜橫切示意圖	100
圖6-2 電漿誘導PHB薄膜接枝聚合丙烯酸反應示意圖	102
圖6-3 PHB薄膜表面接枝聚丙烯酸以EDAC固定幾丁聚醣反應示意圖	103
圖6-4 利用預烘/凍乾法製備非對稱型磷酸鈣/幾丁聚醣複合薄膜之流程圖	104
圖6-5 固定幾丁聚醣之PHB薄膜模具及製備流程示意圖	105
圖6-6 ASTM D638 Type V	107
圖6-7直接冷凍乾燥法製備孔洞型磷酸鈣/幾丁聚醣薄膜	112
圖6-8 50oC預烘20分鐘再冷凍乾燥製備非對稱型磷酸鈣/幾丁聚醣薄膜	113
圖6-9 50oC預烘40分鐘再冷凍乾燥製備非對稱型磷酸鈣/幾丁聚醣薄膜	114
圖6-10不同預烘時間下幾丁聚醣薄膜截面圖,(a)直接凍乾、(b)50oC預烘40分鐘再凍乾與(c)50oC預烘40分鐘再凍乾	115
圖6-11在50oC的不同預烘時間下,磷酸鈣/幾丁聚醣複合薄膜之平衡吸水示意圖	117
圖6-12 直接烘乾的PHB薄膜上表面(a)與經過DMSO處理15分鐘(b)、30分鐘(c)與60分鐘(d)之PHB薄膜的SEM圖	118
圖6-13 將幾丁聚醣與經DMSO處理之PHB薄膜製備成幾丁聚醣/PHB複合薄膜的幾丁聚醣面(a)、PHB面(b)與截面(c)	118
圖6-14電漿處理功率與電漿處理時間對於聚丙烯酸接枝量之影響圖	119
圖6-15 (a)接枝反應之溫度與時間對於丙烯酸接枝量之影響與不同反應溫度(b)60oC、(c)70oC與(d)80oC下反應4小時的PHB薄膜變化	120
圖6-16 (a)未處理之PHB薄膜、(b)接枝聚丙烯酸之PHB薄膜與(c)固定幾丁聚醣之PHB-g-PAA薄膜的FTIR-ATR圖	122
圖6-17 非對稱型幾丁聚醣-PHB複合薄膜(a)幾丁聚醣面、(b)PHB面與(c)截面的SEM圖	123
圖6-18 L929細胞於幾丁聚醣複合薄膜的細胞增生圖	129
圖6-19 L929細胞於非對稱型chitosan/PHB薄膜的細胞增生圖	130
圖7-1 骨瘉合階段示意圖(Frost, 1989a)	134
圖7-2 (a)探討非對稱性薄膜阻隔牙齦纖維母細胞之載具以及(b)細胞實驗配置圖。	138
圖7-3 動物實驗流程	141
圖7-4 牙齦纖維母細胞於非對稱型磷酸鈣/幾丁聚醣複合薄膜上之增生曲線圖,代號說明:chitosan為純幾丁聚醣薄膜,CDHA/chitosan為添加CDHA之幾丁聚醣複合薄膜,BCP/chitosan為添加BCP之幾丁聚醣複合薄膜薄膜,TCP/chitosan為添加TCP之幾丁聚醣複合薄膜,-D為以直接烘乾法所得之緻密薄膜,-P0為以冷凍乾燥法所得之孔洞薄膜,-P20為50oC預烘20分鐘再凍乾之非對稱型薄膜,-P40為50oC預烘40分鐘再凍乾之非對稱型薄膜	144
圖7-5 牙齦纖維母細胞於(a)chitosan-P0與(b)BCP/chitosan-P0上成長不同天數的SEM圖	145
圖7-6 牙齦纖維母細胞於非對稱型幾丁聚醣/PHB複合薄膜的PHB緻密面和幾丁聚醣孔洞面的生長情形,與在幾丁聚醣緻密薄膜之細胞生長情形	146
圖7-7 牙齦纖維母細胞於非對稱型幾丁聚醣/PHB複合薄膜上成長4天後(a)幾丁聚醣面與(b)PHB面上的SEM圖	146
圖7-8 牙齦纖維母細胞於非對稱型磷酸鈣/幾丁聚醣與幾丁聚醣/PHB複合薄膜上培養12周,薄膜下方培養盤之蛋白質濃度變化	148
圖7-9 chitosan-P0與chitosan-P40經過12周培養後,材料之結構變化SEM圖	149
圖7-10 骨母細胞在各種非對稱型磷酸鈣/幾丁聚醣複合薄膜上之細胞增長曲線	151
圖7-11 骨母細胞分別培養在(a)BCP/chitosan-P0與(b)BCP/chitosan-P40孔洞面的SEM圖	151
圖7-11骨母細胞於PHB薄膜、 PHB-g-PAA薄膜、PHB-g-PAA-chitosan薄膜、chitosan/PHB薄膜的孔洞面(chitosan surface)以及BCP-chitosan/PHB薄膜的孔洞面(BCP/chitosan surface)的細胞增生圖	153
圖7-13 經過手術後21天,電腦斷層掃描(micro-CT)重建頭蓋骨缺損恢復型態,1. Black為未覆蓋薄膜之骨修復情形,2. BCP/chitosan-P為覆蓋添加BCP之幾丁聚醣孔洞膜之骨修復情形,3. CDHA/chitosan-P為覆蓋添加CDHA之幾丁聚醣孔洞膜之骨修復情形,4. TCP/chitosan-P為覆蓋添加TCP之幾丁聚醣孔洞膜之骨修復情形,5. chitosan-P為覆蓋純幾丁聚醣孔洞膜之骨修復情形	155
圖7-14 經過手術後21天,以X光拍攝之頭蓋骨影像觀察骨缺損恢復型態,1. Black為未覆蓋薄膜之骨修復情形,2. BCP/chitosan-P為覆蓋添加BCP之幾丁聚醣孔洞膜之骨修復情形,3. CDHA/chitosan-P為覆蓋添加CDHA之幾丁聚醣孔洞膜之骨修復情形,4. TCP/chitosan-P為覆蓋添加TCP之幾丁聚醣孔洞膜之骨修復情形,5. chitosan-P為覆蓋純幾丁聚醣孔洞膜之骨修復情形	156
圖7-15 以螢光顯微鏡觀察bone marker之骨再生區域	158
圖7-16 以硬組織切片觀察未覆蓋薄膜、覆蓋chitosan-P與覆蓋CDHA/chitosan-P的骨缺陷之修復型態,兩箭頭所指位置之間為骨缺損之區域	159
圖7-17 以硬組織切片觀察覆蓋BCP/chitosan-P與覆蓋TCP/chitosan-P的骨缺陷之修復型態,兩箭頭所指位置之間為骨缺損之區域	160
 
表目錄
表2-1 不同結晶型態之磷酸鈣一覽表(Dorozhkin, 2009)	9
表2-2 常用於組織工程之生物可降解合成高分子	10
表4-1 以逆乳化法合成出之CDHA及在800oC與1000oC煆燒所獲得之BCP與TCP之(Ca/P)p,與XRD計算之Ca/P	39
表5-1 常見的黏度常數K與a值	52
表5-2不同類型的薄膜代號	54
表5-3 模擬體液與人體血漿中離子濃度表(Kokubo et al., 1990; Kokubo & Takadama, 2006)	56
表5-4 不同濃度幾丁聚醣溶液通過毛細管之時間與黏度計算一覽表	65
表5-8 磷酸鈣/幾丁聚醣複合薄膜的厚度、上下表面孔徑及孔隙度與總體孔隙度表	70
表6-1 不同類型的薄膜代號	104
表6-2 非對稱型幾丁聚醣複合薄膜的厚度、上下表面孔徑及孔隙度與總體孔隙度	116
表6-3 乾燥狀態下不同幾丁聚醣複合薄膜之機械性質表	126
表6-4 濕潤狀態下不同幾丁聚醣複合薄膜之機械性質表	127
表7-1 以X光計算新生骨的修復面積比例一覽表	157
表7-2 骨缺損以截面積計算新生骨修復面積比例一覽表	161
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