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研究生:陳映達
研究生(外文):Yin-Ta Chen
論文名稱:台灣市售微藻產品之衛生品質以及抗氧化活性探討
論文名稱(外文):Hygienic quality and antioxidant activities of commerical microalgal products in Taiwan
指導教授:蔡永祥蔡永祥引用關係
指導教授(外文):Yung-Hsiang Tsai
學位類別:碩士
校院名稱:大仁科技大學
系所名稱:環境管理研究所
學門:環境保護學門
學類:環境資源學類
論文種類:學術論文
論文出版年:2006
畢業學年度:94
語文別:中文
論文頁數:107
中文關鍵詞:微生物品質一般組成份藻類抗氧化活性抗菌活性
外文關鍵詞:microbiological qualityantioxidant activityproximate compositionantibacterial activityalgae
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本研究係收集市售微藻產品共12個樣品,其中綠藻類與藍綠藻類各有6個樣品,分析測定其化學與微生物品質、抗氧化活性以及抗菌活性。研究結果得知,綠藻製品之平均一般組成為粗蛋白40.78 %,粗脂肪 14.28 %,灰分 7.79 %,水分 6.98 %,碳水化合物 30.17 %;藍綠藻產品之平均一般化學組成為粗蛋白57.36 %,粗脂肪16.57 %,灰分8.07 %,水分7.15 %,碳水化合物10.79 %。然而,除了綠藻製品之碳水化合物含量高於藍綠藻產品外(p<0.05),其他項目並無顯著性差異;但在粗蛋白項目方面如扣除BH–C與BH–S廠牌之膠囊產品,則藍綠藻產品之粗蛋白含量顯著較綠藻製品為高。其次,在微生物品質方面,綠藻與藍綠藻製品各有一件樣品 (分別為4.94與5.30 log CFU/g) 超過食品衛生標準之限量規定 (4.7 log CFU/g),而在所有樣品中均未檢出大腸桿菌(E. coli)與大腸桿菌群(Total coliform)。在總脫鎂葉綠酸鹽含量方面,所有樣品皆低於食品衛生標準之規定(即綠藻類80 mg %與藍藻類100 mg %以下);此外,所有樣品之重金屬含量亦低於食品衛生標準規定之限量;顯示市售食用藻類製品之衛生品質大部分符合相關規定標準。

另外,以市售藻類製品之甲醇萃取物,進行清除??diphenyl-β- picryl-hydrazyl (DPPH)自由基、還原力、羥基自由基清除能力、抑制亞麻油酸自氧化之硫氰酸鐵法、捕捉超氧陰離子能力以及亞鐵離子螯合能力之測定等抗氧化試驗。結果顯示隨著甲醇粗萃物濃度之增加,前述抗氧化試驗等之抗氧化活性會隨之增強,但個體間之抗氧化能力亦有所不同。其中,以清除DPPH自由基、捕捉超氧陰離子與亞鐵離子螯合能力所測定之結果顯示,綠藻與藍綠藻製品間之抗氧化能力相似;但綠藻製品在還原力方面較藍綠藻為佳,其50 %有效濃度(50 % effective concentration, EC50) 分別為3.59與7.10 mg/ml。反之,藍綠藻製品比綠藻製品有較好之羥基自由基清除能力(EC50分別為0.07與0.11 mg/ml);整體而言除了還原力之外,市售微藻甲醇萃取物之抗氧化活性是相當好
。其次,在抗氧化成分分析方面,綠藻與藍綠藻製品中ascorbic acid與β-carotene之含量,除了綠藻製品之GB-D產品β-carotene含量較高為50 mg/g以外,兩者間並無顯著差異;但藍綠藻比綠藻有較高含量之總酚類(平均含量分別為4.7與2.5 mg/g)。再者,綠藻製品之總葉綠素平均含量(24.3 mg/g)較藍綠藻為高(12.2 mg/g),但藍綠藻製品中含有高量之藻藍素平均為29.6 mg/g,而綠藻製品除了GB-D產品含有少量之藻藍素(0.7 mg/g)外,其他產品皆未檢出。

抗菌活性分析方面,以紙錠擴散法初步檢測市售微藻甲醇萃取物之抗菌能力,得知綠藻產品之甲醇萃取物對4株病原菌Staphylococcus aureus、Bacillus cereus和Escherichia coli、Pseudomonas aeruginosa均無抑菌活性。而6個藍綠藻製品中只有2個產品(FE-S與BH-S)具有抑菌之活性,其中FE-S產品甲醇萃取物對於S. aureus之最低抑制濃度(minimum inhibition concentration, MIC) 與最低殺菌濃度(minimum bactericidal concentration, MBC)相同均為25 μg/ml,而對B. cereus之MIC與MBC均為50 μg/ml;其次,BH-S產品甲醇萃取物對S. aureus之最低抑制濃度與最低殺菌濃度相同為125 μg/ml。
Six green algae products and 6 blue-green algae products sold in Taiwanese markets were tested to determine the chemical and microbiological quality, antioxdant activity and antibacterial activity. It was found that the average values of proximate composition in green algae products and blue-green algae products were protein: 40.78 % and 57.36 %, lipid: 14.28 % and 16.57 %, ash: 7.79 % and 8.07 %, moisture: 6.98 % and 7.15 %, carbonhydrate: 30.17 % and 10.79 %, respectively. The average carbonhydrate value of the green algae products was significantly (p < 0.05) higher than that of the blue-green algae products. In contrast, the average protein level in the green algae products was significantly lower than that of the blue-green algae products, except for BH-C and BH-S products. Among them, only one sample of green algae products and one sample of blue-green algae products had aerobic plate count (4.94 and 5.30 log CFU/g, respectively) greater than the Taiwanese regulatory standard limit of 4.70 log CFU/g. None of these samples contained total coliform and E. coli. The amounts of pheophoride a and heavy metals in all samples were below the Taiwanese regulatory level. Almost these tested samples were acceptable products based on Taiwanese regulatory standards with the chemical and microbiological quality in the samples.

The antioxidant properties and antioxidant components of methanolic extracts from microalgal products were studied. Using the scavenging effect on 1,1-diphenyl-2-picryl-hydrazyl radicals, superoxide radicals and chelating effects on ferrous ions, the antioxidant activity of methanolic extracts from green alage products was similar as blue-green algae products. The green algae products showed significantly lower EC50 (50 % effective concentration, EC50) value than blue-green algae products in the reducing power (3.59 and 7.10 mg/ml, respectively). In contrast, the EC50 value of the scavenging effect on hydroxyl radical was lower for blue-green algae products than green algae products (0.07 and 0.11 mg/ml, respectively). Generally, methanolic extracts from green algae and blue-green algae products were good in the antioxidant properties tested. On the other hand, the ascorbic acid and β-carotene contents were similar for both microalgal products, except for GB-D product contained higher β-carotene content (50 mg/g). However, a higher average total phenols level at 4.7 mg/g was found in blue-green algae products than that at 2.5 mg/g in green algae products, whereas the average total chlorophyll content in green algae was higher than that of blue-green algae (24.3 and 12.2 mg/g, respectively). Although the average phycocyanin content at 29.6 mg/g was found in blue-green algae products, only one sample (GB-D product) of green algae products had 0.7 mg/g of phycocyanin.

The methanolic extracts of commercial microalgae products were assayed with disc diffusion method for their antibacterial activity against pathogens: Staphylococcus aureus, Bacillus cereus, Escherichia coli and Pseudomonas aeruginosa. None of the green algae products inhibited the four pathogens. Only 2 of the blue-green algae products (FE-S and BH-S) inhibited the growth of S. aureus and B. cereus. The minimum inhibition concentration (MIC) and minimum bactericidal concentration (MBC) of the methanolic extract from FE-S product against S. aureus was 25 μg/ml and against B. cereus was 50 μg/ml, whereas the MIC and MBC of the methanolic extract from BH-S product against S. aureus was 125 μg/ml.
中文摘要 Ⅰ
英文摘要 Ⅲ
誌謝 Ⅴ
目 錄 Ⅵ
圖目錄 XIV
表目錄 XV
壹、文獻整理 1
一、微藻 1
二、綠藻 2
三、杜氏藻 2
四、螺旋藻 3
五、食用藻類之衛生標準 3
(一)現行食用藻類之衛生標準依行政院衛生署食字第09086640號
公告 3
(三)食品衛生標準之指標菌 3
1.總生菌數 3
2.大腸桿菌群 4
3.大腸桿菌 4
(三)脫鎂葉綠酸鹽( pheophorbide )之生成 6
1. 脫鎂葉綠酸鹽所造成人體之傷害 6
六、自由基的定義 7
七、氧自由基對生物體內所造成之危害 8
八、抗氧化劑與抗氧化機構 9
(一) 抗氧化劑之定義 9
1自由基終止型. 10
2 還原劑或氧清除劑 11
3單重態氧抑制劑. 11
4金屬螯合劑 11
5抗氧化酵素 12
(二) 抗氧化劑之種類 12
1酵素型抗氧化劑. 12
(1) 超氧岐化酵素(superoxide dismutase, SOD) . 12
(2) 麩胱苷肽過氧化酶(glutathione peroxidase, GSHPx) 13
(3) 觸酶 ( catalase ) 13
2非酵素型抗氧化劑. 13
(1) 天然抗氧化劑. 14
A、 維生素E 14
B、 維生素C 15
C、 類胡蘿蔔素 15
D、 葉綠素 16
E、 類黃酮 17
F、 花青素 17
G、 藻藍素 17
(2) 人工抗氧化劑 18
A、 丁基羥基甲氧苯 (butyl hydroxyanisol , BHA ) 18
B、 丁基羥基甲苯(butyl hydroxytoluene, BHT) 18
C、 三級丁氫醌(tert-butyl hydroquinone, TBHQ ) 18
D、 沒食子酸酯 (propyl gallate, PG) 19
九、抗氧化活性測定之原理 19
(一) 硫氰酸鐵法 19
(二) DPPH(ㄐA?diphenyl-β-picrylhydrazyl)自由基清除能力之測
定 20
(三) 還原力之測定 20
(四) 羥基自由基清除能力 21
(五) 捕捉超氧陰離子能力之測定 21
(六) 亞鐵離子螯合能力之測定 22
十、藻類相關之抗菌研究 23
貳、市售微藻製品之化學與微生物品質之探討 35
一、前言 35
二、材料與方法 36
(一) 一般組成分分析 36
1. 水分測定 36
2. 粗蛋白含量測定 36
3. 粗脂肪含量測定 37
4. 灰分 37
5. 碳水化合物 38
(二) 微生物分析 38
1. 總生菌數( Aerobic plate count, APC ) 38
2. 大腸桿菌群( Total coliform )與大腸桿菌( E. coli ) 38
(三) 脫鎂葉綠酸鹽含量 39
(四) 重金屬含量分析 39
(五) 統計分析 40
三、結果與討論 41
(一) 市售微藻之一般化學組成分 41
(二) 市售微藻之微生物與總脫鎂葉綠酸鹽含量 42
(三) 市售微藻之重金屬含量 42
參、市售微藻之抗氧化活性評估 48
一、前言 48
二、材料與方法 50
(一) 微藻產品 50
(二) 抗氧化活性之試藥 50
(三) 抗氧化分析方法 52
1. 微藻產品之萃取(萃取物之製備) 52
2. ??Diphenyl-β-picrylhydrazyl(DPPH)清除能力測定 52
3.還原力之測定 52
4.亞鐵離子螯合能力之測定 53
5. 硫氰酸鐵法 54
6. 清除超氧陰離子能力之測定 55
7. 清除羥基自由基( hydroxyl radical )能力之測定 55
(四) 市售微藻之抗氧化成分分析 56
1. 總酚類化合物含量之測定 56
2. 抗壞血酸之定量分析 56
3. β-胡蘿蔔素含量 57
4. 葉綠素含量之測定 57
5. 藻藍素含量之測定 58
(五) 統計分析 58
三、結果與討論 59
(一) 微藻之萃取 59
(二) 微藻甲醇萃取物之抗氧化活性測定 59
1. DPPH 自由基清除效應 59
2.還原力之測定 60
3.清除羥基自由基之能力 61
4. 清除超氧陰離子(superoxide anion)之能力 62
5. 亞鐵離子螯合能力之測定 63
6. 硫氰酸鐵法測抗氧化能力 64
(三) 市售微藻製品之抗氧化成分分析 66
1. 市售微藻甲醇萃取物之維生素C、β-胡蘿蔔素與總酚
類含量 66
(1) 維生素C含量 66
(2) β-胡蘿蔔素含量 67
(3) 總酚類含量 67
2. 市售微藻產品中葉綠素a、葉綠素b、總葉綠素及藻藍
素之含量 67
(1) 葉綠素a、葉綠素b、總葉綠素含量 67
(2) 藍藻素(phycocyanin)之含量 68
肆、市售微藻之抗菌活性探討 79
一、前言 79
二、材料與方法 81
(一) 微藻產品 81
(二) 實驗菌株 81
(三) 實驗藥品 81
(四) 抗菌活性分析方法 81
1. 微藻甲醇萃取物 81
2. 菌體之活化培養 81
3. 抗菌活性之測試 82
4. 最低抑制濃度(minimum inhibition concentration, MIC)之
測試 82
5. 最低殺菌濃度(minimum bactericidal concentration, MBC)
之測試 82
三、結果與討論 84
(一) 市售微藻抗菌活性之測試 84
(二) 最低抑制濃度(MIC)測試 84
(三) 最低致死濃度(MBC)測試 84
伍、結論 90
陸、參考文獻 92
附錄 107
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