臺灣博碩士論文加值系統

中文摘要

台灣曾發生因食用海苔產生光過敏的罕見案例，本實驗室分析得知致毒物質為葉綠素衍生物焦脫鎂葉綠酸鹽（pyropheophorbide a）。為知可食性海藻之食用安全性，故探討台灣常見的三種可食性藻類紫菜、龍鬚菜與石蓴經儲藏及乾濕熱加工後對葉綠素類物質之影響，其次粗萃葉綠素酶並探討生化特性，最後分析市售海藻類乾製品葉綠素類物質（chlorophyll a compounds）含量與色澤。
經檢測三種生鮮藻類chlorophyll a含量隨儲藏天數增加而下降，在3℃溫度下平均下降70.67﹪，而在25℃下則下降95.74﹪。在20℃～100℃的乾燥後發現，在20℃下乾燥，三種藻類chlorophyll a降解緩慢且降解產物含量低。縐紫菜於60℃下乾燥，其chlorophyll a降解快，降解產物在各溫度下含量均未超過1400 ppm。龍鬚菜chlorophyll a於60℃下乾燥2小時後降至總量的28﹪。石蓴pheophytin a含量於40℃下乾燥16小時升高至2105 ppm。市售復水紫菜於20℃下乾燥16小時後並無檢測到pyropheophorbide a，但40℃下乾燥2小時使pyropheophorbide a含量達658 ppm。另外，進行溼熱加工後三者pyropheophorbide a於10分鐘後均無法測得。
縐紫菜與龍鬚菜葉綠素酶的最適作用溫度為50℃，石蓴者為40℃。縐紫菜葉綠素酶最適作用pH值為 8.0，龍鬚菜為pH 9.0，石蓴為pH 7.0。縐紫菜與龍鬚菜葉綠素酶在20～50℃之間可持續保留完整活性；石蓴chlorophyllase活性在20～40℃保持完整。三者葉綠素酶均對chlorophyll a有較高的催化活性。
檢測台北、基隆、台中與高雄地區之市售海藻類乾製品中葉綠素類物質含量，由結果得知，高雄地區之乾紫菜與海苔片檢體在pyropheophorbide a含量上高於台北地區者；各地區乾紫菜與海苔片之產品種類在葉綠素類物質含量上無明顯差異。在Hunter Lab值方面，台北地區乾紫菜檢體之a值與其海苔檢體者呈現顯著差異；高雄地區乾紫菜與其海苔檢體在彩度與色相角上呈現顯著差異。市售海藻類乾製品中L值只在台北地區和基隆地區之海苔片檢體中有地域性差異。

Abstract

An infrequent photosensitization case due to ingesting dried laver flakes had happened in Northern Taiwan. Our previous research suggested that the causative substances were chlorophyll a derivatives, pheophorbide a and pyropheophorbide a. The research is to study the temperature effects of storage and heat process on the content of pheophorbide a and pyropheophorbide a in three kinds of fresh edible seaweeds, Porphyra crispata, Gracilaria tenuistipitata and Ulva lactuca and one kind of market semi-processed laver, and then we invesgated the characteristics of chlorophyllase in the fresh edible seaweeds. Moreover, we will also investigate the levels of chlorophyll a derivatives and Hunter Lab values in market dried laver products.
The chlorophyll a contents of fresh seaweeds caused 70.67﹪and 95.74﹪losses during the storage process in 3℃ and 25℃, respectively. After dry heating at 20℃, the losses of chlorophyll a in seaweed samples were much slower and the degradative derivatives were less. When P. crispata heating at 60℃, the levels of chlorophyll a decreased faster than other heating temperature, and the levels of degradative derivatives were lower than 1400 ppm. The levels of chlorophyll a of G. tenuistipitata decreased to 28﹪after heating at 60℃for 2 hr. The pheophytin a contents of U. lactuca increased to 2105 ppm after heating at 40℃ for 16 hr. And the levels of pyropheophorbide a of market semi-processed laver were non-detectable after heating at 20℃ for 16 hr, but did cause its increase to 658 ppm when heating at 40℃ for 2 hr. In the study of the effect of blanching on the chlorophyll a compounds of three kinds of fresh seaweeds in boiling water, the levels of pyropheophorbide a were not detected after dipping in boiling water for 10 min.
The temperatures of maximum enzymatic activity of chlorophyllase were 50℃ for P. crispata and G. tenuistipitata, and 40℃ for U. lactuca. The optimum pH for P. crispata, G. tenuistipitata and U. lactuca were pH 8.0, 9.0 and 7.0, respectively. The chlorophyllase showed a considerable resistence to thermal treatment. The enzyme was stable for incubating at 50℃ for 30 min. And the activity was higher with substrate chlorophyll a than with pheophytin a.
Finally, we analyzed 40 samples of commercial seaweed products (dried lavers and dried laver flakes) sold in markets of Taipei, Keelung, Taichung and Kaohsiung. The results indicated the levels of pyropheophobide a of both dried lavers and dried laver flakes in Kaohsiung were higher than those in Taipei. In the aspect of Hunter Lab values between dried lavers and dried laver flakes, the a value was significantly different in Taipei’s samples, but the chroma and the hue angle were noted in Kaohsiung’s samples. Meanwhile, the L value of dried laver flakes collected from Taipei and Keelung was significantly varied.

目 錄

中文摘要 ‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧1
英文摘要 ‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧3

第一章 研究背景與目的 ‧‧‧‧‧‧‧‧‧‧‧‧‧‧5
壹、研究背景‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧6
貳、研究目的‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧8

第二章 文獻回顧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧10
壹、海藻的定義與重要性‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧11
貳、紫菜的種類與加工利用‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧15
參、葉綠素簡介‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧17
肆、葉綠素於光合性生物中的降解‧‧‧‧‧‧‧‧‧‧‧‧‧‧18
伍、葉綠素於加工過程中的降解‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧20
陸、葉綠素酶的分布及催化反應‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧22
柒、葉綠素酶的生化特性‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧24

第三章　研究內容‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧26
壹、縐紫菜、龍鬚菜與石蓴經乾熱、溼熱加工及儲藏程序後葉綠素
及衍生物之變化‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧27
一、前言‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧27
二、實驗材料與藥品‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧27
（一）實驗材料‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧27
（二）實驗試藥‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧28
三、實驗方法‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧28
（一）儲藏試驗‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧28
（二）乾熱加工及濕熱加工‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧28
（三）葉綠素類物質之萃取‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧29
（四） HPLC分析方法‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧29
（五）標準品之檢量線製作‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧30
四、結果‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧32
（一）儲藏溫度對三種生鮮海藻中葉綠素降解之影響‧‧‧‧32
（二）乾熱加工對三種生鮮海藻中葉綠素降解之影響‧‧‧‧33
（三）濕熱加工對三種生鮮海藻中葉綠素降解之影響‧‧‧‧35
五、討論‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧36
（一）藻類葉綠素及其降解產物之萃取與分析‧‧‧‧‧‧‧36
（二）葉綠素在儲藏期間的降解及衍生物的生成‧‧‧‧‧‧37
（三）葉綠素在乾熱加工過程中的降解及衍生物的生成‧‧‧38
（四）葉綠素在濕熱加工過程中的降解及衍生物的生成‧‧‧41
圖表‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧42

貳、縐紫菜、龍鬚菜與石蓴葉綠素酶特性分析‧‧‧‧‧‧‧‧‧53
一、前言‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧53
二、實驗材料與藥品‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧53
（一）實驗材料‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧53
（二）實驗試藥‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧53
三、實驗方法‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧54
（一）萃取催化受質chlorophyll a ‧‧‧‧‧‧‧‧‧‧‧‧54
（二）純化催化受質chlorophyll a ‧‧‧‧‧‧‧‧‧‧‧‧55
（三）製備葉綠素酶丙酮粉‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧56
（四）製備葉綠素酶粗酵素液‧‧‧‧‧‧‧‧‧‧‧‧‧‧56
（五）葉綠素酶蛋白質濃度測定‧‧‧‧‧‧‧‧‧‧‧‧‧57
（六）葉綠素酶活性測定‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧57
（七）最適pH值的測定‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧58
（八）最適溫度的測定‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧59
（九）酵素之高溫耐受性的測定‧‧‧‧‧‧‧‧‧‧‧‧‧59
（十）酵素對受質chlorophyll a與pheophytin a之
比活性與親和性‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧59
四、結果‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧60
（一）葉綠素酶催化受質chlorophyll a的製備‧‧‧‧‧‧‧60
（二）縐紫菜、龍鬚菜及石蓴葉綠素酶之最適溫度‧‧‧‧‧60
（三）縐紫菜、龍鬚菜及石蓴葉綠素酶之最適pH值‧‧‧‧61
（四）縐紫菜、龍鬚菜及石蓴葉綠素酶之高溫耐受性‧‧‧‧62
（五）縐紫菜、龍鬚菜及石蓴葉綠素酶對chlorophyll a
與pheophytin a的親和性與比活性‧‧‧‧‧‧‧‧‧62
五、討論‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧63
（一）縐紫菜、龍鬚菜及石蓴葉綠素酶之最適溫度‧‧‧‧‧ 63
（二）縐紫菜、龍鬚菜及石蓴葉綠素酶之最適pH值‧‧‧‧63
（三）縐紫菜、龍鬚菜及石蓴葉綠素酶之高溫耐受性‧‧‧‧64
（四）縐紫菜、龍鬚菜及石蓴葉綠素酶之基質親和性‧‧‧‧64
圖表‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧66



參、市售海藻類乾製品中葉綠素類物質之調查‧‧‧‧‧‧‧‧‧70
一、前言‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧70
二、實驗材料與藥品‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧70
（一）實驗材料‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧70
（二）實驗試藥‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧70
三、實驗方法‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧71
（一）量測市售海藻類乾製品之Hunter Lab值‧‧‧‧‧‧‧71
（二）萃取市售海藻類乾製品中的葉綠素類物質‧‧‧‧‧‧71
（三）統計分析‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧71
四、結果‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧72
（一）市售海藻類乾製品中葉綠素類物質的含量‧‧‧‧‧‧72
（二）市售海藻類乾製品之色澤測定‧‧‧‧‧‧‧‧‧‧‧72
五、討論‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧74
（一）市售海藻類乾製品中葉綠素類物質的含量‧‧‧‧‧‧74
（二）綠色度的量測‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧75
（三）市售海藻類乾製品之色澤與葉綠素類物質含量
的相關性‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧75
圖表‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧77

參考文獻‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧88
附錄‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧101
謝誌‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧106

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