台灣原生種土肉桂葉中類黃酮糖?分離純化製程之研究探討

Study on isolation processes of bioactive flavonol glycosides from Taiwan endemic plant Cinnamomum osmophloeum leaves

黃偉健(Wei-chien Husng)

指導教授 ： 詹效松 ； 曾耀銘

朝陽科技大學/理工學院/應用化學系碩士在職專班/碩士(2010年)

摘要

台灣原生種土肉桂(Cinnamomum osmophloeum Kaneh)為台灣特有之樟科喬木，生長於海拔400-1500 m的天然闊葉林中，台灣土肉桂葉傳統民間用藥用於治療發炎、腸道感染、糖尿病等，先前本實驗室研究小組以報告類黃酮醣?(kaempferol glycoside)已有抗發炎、糖尿病等生理活性，所以本研究使用毛細管電泳定量方法，以兒茶素為內標準品分析土肉桂葉中三種類黃酮醣?kaempferitrin (C1), kaempferol3-O-β-D-glucopyranosyl-(1-4)-α-L-rha mnopyranosyl-7-O-α-L–rhamnopyranoside (C2), and kaempferol3-O-β-D-apio furanosyl-(1-2) –α -L-arabinofuranosyl-7-O- α-L-rhamnopyranoside (C3)，毛細管電泳分析最佳條件為四硼酸納鹽緩衝溶液濃度35 mM， pH 9.8，分離電壓25 kV，樣品注射時間5 s，溫度25 oC，每一種化合物濃度範圍0.075–0.6 mg/mL (C1, C2 and C3)來進行分析，此方法已建立應用於分析土肉桂葉中三種類黃酮醣?(C1, C2 and C3)。另一方面，我們也開發了以鹽水(0.1 M NaCl)萃取系統來提取土肉桂葉中三種類黃酮醣?C1，C2和C3，我們將乾燥後的土肉桂葉在室溫下以0.1 M NaCl溶液萃取2小時，萃取液經由過濾後，分成兩部份，程序A是將以電氣透析儀脫鹽後的萃取液導入聚合離子交換樹脂HP-20進行吸附、脫附以回收C1，C2和C3，程序B是將我們過濾後的萃取液不經由電氣透析儀脫鹽直接導入聚合離子交換樹脂HP-20進行吸附、脫附以回收C1，C2和C3，上述過程中將HP-20樹脂置入於玻璃管柱中(L 31 cm × OD 3.3 cm)其樹脂填充量為36g，在脫附試驗中是以95 %乙醇作為脫附劑，在程序A中C1，C2和C3的回收率分別為 95.36 %， 92.21 %和96.39 %，而程序B的回收率分別為，94.60 %，94.72 %和97.81 %，以毛細管電泳分析兩個流程相對含量的對比。

關鍵字

HP-20 ； 毛細管電泳 ； 定量分析 ； 台灣原生種土肉桂

並列摘要

Cinnamomum osmophloeum Kaneh. (Lauraceae) is an indigenous tree that grows in Taiwan’s natural hart wood forest at an elevation between 400-1500 m. Traditionally, the leaves of C. osmophloeum are used in folk medicine for the treatment of inflammation, intestinal infections, astringent, diuretic and diabetic complications in Taiwan. Previously our research group have been reported the anti-inflammatory and anti-diabetic flavonoid (kaempferol glycosides) from the leaves of this species. In the present study, a capillary electrophoresis quantification method, using catechin as the internal standard was developed for the analysis of kaempferitrin (C1), kaempferol 3-O-β-D-glucopyranosyl -(1-4)-α-L-rhamnopyranosyl-7-O-α-L-rhamnopyranoside (C2), and kaem pferol 3-O-β-D-apiofuranosyl-(1-2)-α-L-arabinofuranosyl-7-O-α-L-rham nopyranoside (C3). Optimal conditions found were 35 mM sodium borate buffer, pH 9.8, separation voltage 25 kV, hydrodynamic injection time 5s and temperature 25 ºC. Linearity was found over the 0.075–0.6 mg/mL concentration ranges of each compound (C1, C2 and C3) analyzed. The developed method has been applied for determination of C1, C2 and C3 in a series of C. osmophloeum leaves. On the other hand, we also developed a simple extraction process using salt water (0.1 M NaCl) for the separation of C1, C2 and C3 from C. osmophloeum leaves. The air-dried powder of C. osmophloeum leaves was extracted for 2 h with 0.1 M NaCl under stirring at room temperature. The extract was filtered and separated into two parts. In the first part the NaCl was removed using electro dialysis and subsequently introduced into a macroporous adsorption resin, where as the second part directly (without NaCl remove) introduced to adsorption resin for recovering the C1, C2 and C3. The adsorption was performed on Diaion HP-20 resin packed in a glass column (L 31 cm × OD 3.3 cm, with 36 g resin), and desorption was carried out using 95% ethanol. Through the process, in the first part the recoveries of C1, C2, and C3 were 95.36%, 92.21%, and 96.39%; whereas the same in the second part were 94.60%, 94.72%, and 97.81%, respectively. The content of C1, C2, and C3 in the above both parts were quantitatively determined using capillary electrophoresis and found the results was comparable.

並列關鍵字

Cinnamomum osmophloeum ； Diaion HP20 ； capillary electrophoresis ； quantitative analysis

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