青剛櫟之細胞培養株中多酚性成分分析方法的探討
中文摘要
青剛櫟普遍生長在台灣 800 公尺以下之低海拔及平地地區，其新鮮葉中多酚 性成分含量豐富，為了方便探討不同季節、地區間青剛櫟之遺傳變異大小和 變異型式，以利選種、育林及復育之依據、醫療資源之開發，本實驗擬以青 剛櫟新鮮的葉片、枝條、種子及其細胞培養株的萃取液，建立以高效液相層 析法分析其多酚性成分，達到簡捷、穩定、精確的分析條件。
本實驗應用 80% 丙酮為萃取液，萃取青剛櫟及其細胞培養株中之多酚性成分，萃取液經濃縮後，先以二氯甲烷洗去低極性物質，再以通過 C18 cartridge 之方式濾除醣類物質，最後使用逆向層析管柱、酸性的梯度移動相條件及高 感度 photodiode-array 偵測器之運用，得到有效且再現性良好的分析條件。
本次實驗結果，高效液相層析分析方法方面， r2 均在 0.999 以上，最低可偵 測濃度 (LOD, limit of detection) 在 2.5 ~ 10μM 間，最低可定量濃度 (LOQ, lim it of quantity) 在 5 ~ 12.5μM 間。從葉片中檢出 (+)-catechin 、 (-)-epicatechin、 (+)-gallocatechin 、 (-)-epigallocatechin 、 procyanidin-B-4 及 rutin ；枝條及 種子則檢出 (+)-catechin 、 (-)-epicatechin 、 gallic acid 及少量之 (+)-gallocatec hin ；細胞培養株中之多酚性成分多數可測出 (+)-catechin 、 (+)-gallocatechin 及 gallic acid 。
Studies on the analytical method for polyphenols from cell culture lines of Cyclobalanopsis glauca
英文摘要
Cyclobalanopsis glauca (Thunb. ex Murray) Oerst. are distributed very commen in forests fro m sea level to 800m throughout Taiwan island. They are rich in the polyphenolic compounds i n their leaves. To study the polyphenolic components of Cyclobalanopsis glauca in different re ason or area, we developed an analytical method using high-performance liquid chromatograp hy to separate the polyphenolic compounds in the leaves, branches, seeds and it’s cell culture l ines from Cyclobalanopsis glauca.
An aqueous acetone (80%) were added to extract the polyphenols in Cyclobalanopsis glauca a nd it’s cell culture lines. After condensing, the extract was delipidized with dichloromethane a nd pass through C18 cartridge to filter out the sugars in the samples. Then it was chromatogra phed on reversed-phase column with acidic gradient mobile phase system and detected with p hotodiode-array. We found a well-defined, reproducible system for the separation of such poly phenols in the samples.
The results shows the value of r2 for the analysis is above 0.999. The limit of detection (LOD) is in the range of 2.5 ~ 10 μM, the limit of quantity (LOQ) is 5 ~ 12.5μM. In the samples of le aves, there are (+)-catechin, (-)-epicatechin, (+)-gallocatechin, (-)-epigallocatechin, procyanidi n-B-4 and rutin were detected. About branches and seeds, we found (+)-catechin, (-)-epicatech in and gallic acid. Most of the cell culture lines, (+)-catechin, (-)-epicatechin, (+)-gallocatechin were identified.
