以DL-樟脑-10-磺酸为离子对试剂反相高效液相色谱测定血浆中小檗碱的含量

DL-Camphor-10-Sulfonic Acid as an Ion-Pair Reagent for Determination of Berberine in Plasma by Reversed-Phase High Performance Liquid Chromato-graphy

小檗碱(Berberine)属季铵型异喹啉类生物碱。系黄连、黄柏、三颗针等中草药的主要有效成分,具有抗菌消炎作用。近年来发现小檗碱具有抗心律失常和正性肌力等心血管药理作用。生物样品中小檗碱的测定方法有纸色谱法,纸色谱-分光光度法,同位素标记法,气相色谱-质谱法。DL-樟脑-10-磺酸(CSA)是一种手性离子对试剂,曾用于生物胺的测定取得了良好的结果。我们用CSA作离子对试剂,建立了血浆中小檗碱的反相离子对色谱测定方法,并对GSA的色谱行为及方法的优化作了探讨。

Plasma (0. 2ml) was mixed with acetonitrile ( 0. 4ml) containing 13-methylberbere (0. 9ng/ml; internal standard) and centrifuged at 19000g for 10 min. The supernatent (0. 2ml) was mixed with equal volume of mobile phase and centrifuged at 19000g for 5min. The supernatant solution (100uμl) was analysed by HPLC on Nova-Pak C18(4μm) column (15cm×3. 9mm) with acetone, water, dl-camphor-10-sulfonic acid, diethyl amine, acetic acid (20 : 80 : 0. 464 : 0. 5 : 0. 7) as mobile phase (pH 4. 8, Flow 1ml/min), and detected at 345nm. The calibration graphs were rectilinear from 0. 272 to 4. 354 μg/ml plasma. The detection limit of berberine was 4. 5ng. The coefficient of variation of within-day and between-day were 2. 00% (n = 6) and 2. 56% (n = 4) respectively. Mean revoveries were 99.94%.