臺灣博碩士論文加值系統

吳郭魚 (Oreochromis aurea) 皮的一般成分組成水分、粗蛋白、粗脂肪及灰分，分別為 33.38%、32.23%、4.21% 及 0.21%。吳郭魚皮以 0.5 M 醋酸分別在室溫 (20 ± 2℃) 及 50℃ 萃取 2-8 小時，可萃取出酸可溶膠原蛋白 (acid - soluble collagen, ASC)。室溫萃取所得 ASC (ASC-RT) 及 50℃萃取所得之 ASC (ASC-50℃) 之黏度及固型物含量 (Brix) 隨萃取時間的增加而上升，ASC-50℃ 之 Brix 由 3.4 升至 3.6 %，ASC-RT 由 3.0 升至 3.2%。吳郭魚皮以室溫萃取 6 小時可得最大黏度達 777.67 cP，若於 50℃ 萃取 2 小時至 8 小時，黏度由 12.98 cP 些微降至 10.00 cP。吳郭魚在室溫萃取2 小時至 8 小時，所得之 ASC 乾重產率由 19.28 % 增至 28.71%，經二次重複萃取產率可增至 30%；若以 50 ℃ 萃取 2 至 8 小時，產率則無顯著差異 (P>0.05)。ASC-RT 之分子量分布由 105 kD 至 295 kD；ASC-50℃ 之分子量分布為 71 kD 至 244 kD。ASC-RT 最大分子量為 295 kD，濃度隨著萃取時間的增加，由 1.67 降至 1.54 mg/ ml albumin；ASC-50℃ 最大之分子量為 244 kD，隨著萃取時間 2 小時增加至 8 小時，濃度由 2.12 降至 1.53，相反的，分子量為 71 kD 之蛋白當萃取時間 由 2 小時增加至 8 小時，濃度由 1.45 增至 1.56。本研究之 ASC 分子量皆高於市售產品 (MW ≦50 kD) 及魚鱗膠原胜肽 (1.3 - 4.5 kD)，以膠原蛋白酶水解膠原蛋白，可得較小之分子量產物，約為 35 kD 及小於 20 kD。吳郭魚皮萃取 2 小時所得 ASC-RT 較 ASC-50℃ 含較多羥脯氨酸及羥離胺酸，故 ASC-RT 結構可能較 ASC-50℃ 穩定，因此 ASC-RT 的分子量高於ASC-50℃。ASC-RT 及 ASC-50℃ 凍結乾燥後以顯微鏡觀察，兩者結構皆有膠原蛋白纖維存在，以 ASC-50℃ 之結構較 ASC-RT 更纖細且較少孔洞。

The proximate composition of Oreochromis niloticus skin was analyzed. The contents of moisture, crude protein, lipid and ash of tilapia skin were 33.38%, 32.23%, 4.21% and 0.21%, respectively. Tilapia skin collagen was extracted with 0.5 M acetic acid at room temperature (20 ± 2℃) and 50℃ for 2 to 8 h to obtain acid - soluble collagen (ASC). The viscosity and Brix of room-tempeature extracted and 50℃-extracted ASC increased with extraction time. ASC hydrolyzed at room temperature had lower Brix than that at 50℃. Brix for ASC extracted at 50℃ (ASC-50℃) was from 3.4 to 3.6% and ASC extracted at room temperature (ASC-RT) was from 3.0 to 3.2% Brix. The viscosity of ASC solution was maximal at 777.67 cP after 6 h of extraction at room temperature. Conversely, the viscosity of collagen solution extracted for 2 h to 8 h from tilapia skin at 50℃ decreased slightly from 12.98 cP to 10.00 cP. The yield of acid-soluble collagen extracted at room temperature from 2 to 8 h increased from 19.28 % to 28.71% (on dry weight basis), respectively. The yield increased to about 30% after re-extraction. Differences in the yields of extraction at 50℃ from 2 to 8 h were insignificant (P>0.05). The molecular weight of ASC-RT ranged 105 kD to 295 kD, while that extracted at 50℃ had molecular distribution from 71 kD to 244 kD. The largest molecular weight band (295 kD) of ASC-RT decreased in concentration (mg/ ml albumin) from 1.67 to 1.54 mg/ ml. Similarly, concentration of ASC-50℃ of the largest molecular weight band (244 kD) decreased from 2.12 mg/ ml for 2 h and 1.53 mg/ ml for 8 h while the concentration of 71 kD showed an increased trend, enhancing from 1.45 to 1.56 mg/ml for 2 h and 8 h, respectively. The molecular weight (MW) of ASC in present research is much higher than those in commercial products and fish-scale collagen peptides, MW ≤ 50 kD for commercial products and 1.3 - 4.5 kD for fish-scale collagen peptides. Collagen hydrolyzed by collagenase had low molecular weight about 35 kD and less than 20 kD. ASC-RT of 2 h extraction might be more stable in structure than ASC-50℃due to higher hydroxyproline and hydroxylysine content in ASC-RT. Therefore, the molecular weight of ASC-RT was higher than that of ASC-50℃. Microscopic images of ASC-RT and ASC-50℃ were taken to compare differences in ASC microstructure. It seemed that microstructure of freeze-dried ASC-50℃ was fine and less porous than that of ASC-RT. There were presence of collagen fibers with thin strands and collagen sheets in both freeze-dried ASC-RT and ASC- 50℃.

TABLE OF CONTENTS i
ACKNOWLEDGEMENT iv
ABSTRACT v
LIST OF TABLES vii
LIST OF FIGURES viii
1. INTRODUCTION 1
2. LITERATURE REVIEW 3
2.1. Composition and structure of collagen 3
2.2. Fish collagen types 4
2.3. Thermal denaturation temperature of fish collagen 6
2.4. Differences between fish and mammalian collagen 7
2.5. Differences of collagen, gelatin and collagen hydrolysate 8
3. EXPERIMENTAL DESIGN 9
4. MATERIALS AND METHODS 11
4.1. Materials 11
4.1.1 Preparation of fish skin 11
4.1.2 Preparation of acid-soluble collagen 11
4.2. Methods 12
4.2.1 Proximate compositions of tilapia skin 12
4.2.2 Determination of viscosity and Brix of collagen solution 15
4.2.3 Sodium dodecyl sulfate-polyacrylamide gel electrophoresis 15
4.2.4 Molecular weight and concentration determination of commercial products 16
4.2.5 Collagenase hydrolysis of the acid soluble collagen 17
4.2.6 Amino acid analysis 18
4.2.7 Microscopic examination 18
4.2.8 Statistical analysis 19
5. RESULTS AND DISCUSSION 20
5.1. Differences in proximate composition of Taiwan tilapia skin from other fish skin 20
5.2. Physical properties of acid-soluble collagen solution 21
5.2.1. Viscosity and Brix of extracting solution 21
5.2.2. Yields of acid-soluble collagen 24
5.3. Molecular weight distribution of the acid-soluble collagen (ASC) affected by hydrolysis time 27
5.3.1. Electrophoretic pattern of acid-soluble collagen extracted from tilapia skin using different concentration of separating gel. 27
5.3.2. Molecular weight distribution of acid-soluble collagen (ASC) extracted at room temperature and 50 ℃ 29
5.3.3. The molecular weight range of commercial products 33
5.4. Reduction of low molecular weight ASC by collagenase hydrolysis 37
5.5. Hydroxyproline and proline content of acid-soluble collagen 40
5.6. Microstructure of lyophilized acid-soluble collagen 43
6. CONCLUSIONS 46
REFERENCES 47

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