玄參藥材中高極性成分之分離及高效液相層析定量研究   全文下載

Isolation and Determination by HPLC of Polar Constituents from Scrophularia ningpoensis

本研究選擇玄參藥材中高極性，可使用高效液相層析分析，在紫外線波長278nm具有吸收之成分，探討分離新方法。分離純化而得之2-（3-hydroxy-4-methoxyphenyl） ethyl 1-O-[α-L-arabinopyranosyl（1→6）]-feruloyl（1→4）]-α-L-rhamnopyranosyl（1→3）-β-D-glucopyranoside （SN-A），harpagoside（SN-B）及cinnamic acid（SN-C）三成分作為指標成分，利用高效液相層析儀探討其定量方法，並調查市售玄參藥材之指標成分含量。高效液相層析法使用逆相層析管柱（Cosmosil 5C18-AR），移動相為乙?及1.0%醋酸溶液，採線性梯度沖提[15：85→ 27：73 （44 min） ]，以SN-A（20～150 μg／ml），B（5～50μg／ml）及C（20～200μg／ml）做為對照用標準品。其線性迴歸方程式及相關係數（r）分別為SN-A ： Y=14.58X+11.25 （r=0.9907）；SN-B：Y=2.26X－0.40 （r=0.9994）；SN-C：Y=14.12X－29.28 （r= 0.9981），均呈良好線性關係。三種指標成分之回收率，均在100±4%之內。同日內及異日間相對標準偏差試驗，其同日內為0.93～1.01%，異日間為0.28～1.97%，顯示高效液相層析法用於玄參指標成分之分析，具良好之再現性及準確性。測定市售玄參藥材中三種指標成分之含量。結果SN-A之含量平均為0.30%（0.05%～1.71%），含量高低相差34倍；SN-B之含量平均為0.06%（未檢出～0.17%）；SN-C之含量平均為0.25%（0.06%～0.53%），含量高低相差9倍。顯見各成分之含量分布範圍甚大，以SN-A相差34倍最高，而有一件藥材未檢測出SN-B成分。

Three polar constituents, 2-(3-hydroxy-4-methoxyphenyl) ethyl 1-O-[α-L-arabinopyranosyl (1→6)]-feruloyl(1→4)]-α-L-rhamnopyranosy I (1→3)-β- D- glucopyranoside(SN-A), harpagoside(SN-B) and cinnamic acid(SN-C) have been isolated from Scrophularia ningpoensis by a modified isolation method. These constituents were used as marker substances for quality analysis of Scrophulariae Radix. In this study we have also developed a high performance liquid chromatographic method for simultaneous determination of above marker substances in Scrophulariae Radix. This performance was done by using a gradient elution with a solution of acetonitrile in 1.0% acetic acid on a ODS column. The linear calibration ranges were SN-A 20- 150µg/ml (r=0.9907), SN-B 5-50µg/ml (r=0.9994) and SN-C 20-200µg/ml (r=0.9981). Recoveries were SN-A 100.47 ± 3.23%, SN-B 98.49 ± 2.62% and SN-C 96.15 ± 4.39%. The relative standard deviations of three marker substances for intraday and interday analyses were 0.93-1.01% and 0.28-1.97%.. The contents of three marker substances in 24 crude drugs of Scrophulariae Radix were SN-A 0.05%~1.71%, SN-B non detected~0.17%, and SN-C 0.06%~0.53%, respectively.