LC-MS/MS测定6种丹参主要有效成分及其脑靶向分布研究

Simultaneous determination of six salvia miltiorrhiza gradients in rat plasma and brain by LC-MS /MS

建立测定大鼠血浆及脑匀浆中原儿茶酸、原儿茶醛、丹酚酸A、丹酚酸B、隐丹参酮、丹参酮Ⅱ。的LC—MS／MS分析方法。实验采用液液萃取的方法对血浆及脑匀浆样品进行处理，采用Agilent Eclipse Plus—C，。色谱柱（2．1mm×50mm，3．5μm），以0．1％甲酸乙腈和0．1％甲酸水为流动相进行梯度洗脱。质谱条件为电喷雾电离源（ESI），多反应监测（MRM）。正离子模式下检测的离子对（m／z）为297．4／254．3（隐丹参酮），295．5／249．3（丹参酮11。），285．2／154．0（内标地西泮）；负离子模式下检测的离子对（m／z）为154．3／153．1（原儿茶酸），137．3／108（原儿茶醛），493．0／295．2（丹酚酸A），718．0／520．0（丹酚酸B），321．4／152．3（内标氯霉素）。血浆及脑匀浆中隐丹参酮、丹参酮Ⅱ。、原儿茶酸、原儿茶醛、丹酚酸A、丹酚酸B的线性范围分别为：0．15—1000，0．625～1000，0．625～1000，0．625～1000，1．25—1000，2．5—1000μg·L^-1；定量下限分别为0．15，0．625，0．625，0．625，1．25，2．5μg·L^-1建立的方法稳定、灵敏，可用于研究丹参6种成分的脑及血浆药物代谢动力学特性。通过动物实验比较了6种成分之间的血脑分配系数，初步评价其跨越血脑屏障的能力。

To develop a LC-MS/MS method for the determination of protocatechuic acid, protocatechuic aldehyde, salvianolic acid A, salvianolic acid B, cryptotanshinone and tanshinone U A in rat plasma and brain. The plasma and brain samples were precipitated with ethyl acetate, then were separated on an Agilent eclipse plus-C18 column （2. 1 mm × 50 mm, 3.5 μm） using acetonitrile （consisting of 0. 1% formic acid） and water （consisting of 0. 1% formic acid） as mobile phase in gradient elution mode. The mass spectrometer was operated under both positive and negative ion mode with the ESI source, and the detection was performed by MRM. The transition of 154. 3/153. 1 m/z for protocatechuic acid, 137.3/108 m/z for protocatechuic aldehyde, 493.0/295.2 m/z for Salvi- anolic acid A, 718.0/520.0 m/z for salvianolic acid B, 321.4/152. 3 m/z for chloramphenicol, 297.4/254. 3 m/z for cryptotanshi- none, 295.5/249. 3 m/z for tanshinone Ⅱ A and 285.2/154. 0 m/z for Diazepam. The calibration curves in the range of 0. 625-1 000 μg ·L^-1 for protocatechuic acid and protocatechuic aldehyde, 1.25-1 000μg·L^-1 for salvianolic acid A, 2. 5-1 000 μg·L^-1 for salvianolic acid B, 0. 15-1 000μg·L^-1 for cryptotanshinone, 0. 625-1 000 μg·L^-1for tanshinone I1 A are with good linearity in rat plasma and brain. The analysis method is sensitive, simple, and suitable enough to be applied in the pharmacokinetic study of the 6 main components. Animal testing gives the lgBB of the drugs and further studies of the 6 components cross the blood-brain barrier can be carried out.