LC-MS/MS法测定浸润麻醉豚鼠血浆中罗哌卡因浓度

Determination of Ropivacaine in Guinea Pig Plasma after Infiltration Anaesthesia by Liquid Chromatography-tandem Mass Spectrometry

目的建立测定豚鼠血浆中罗哌卡因浓度的液相三重四极杆串联质谱（LC-MS／MS）法，用以测定豚鼠局部浸润麻醉皮内注射后血浆中罗哌卡因浓度，并考察罗哌卡因药动学。方法血浆样品用蛋白沉淀法进行预处理后HPLC-MS／MS法测定罗哌卡因含量。色谱系统：ZORBAXSB-C18（3．0mm×150mm，3．5μm），甲醇-水（80：20，内含5mmol·L^-1甲酸铵）为流动相，流速0．4mL·min^-1，柱温25℃。通过电喷雾离子源（ESI）三重四极杆串联质谱，在正离子模式下以多重反应选择离子检测（MRM）。色谱分析时间为4min。药动学研究中豚鼠皮内浸润麻醉给药0．5％罗哌卡因注射液，按时测定血浆药物浓度。采用TOPFIT软件计算药动学参数。结果罗哌卡因在6．58～6580ng·mL^-1内线性良好（r=0．9991），方法的最低检测浓度为1ng·mL^-1，方法回收率在95．6％～96．6％之间，日内、日间精密度（RSD）均小于6％。豚鼠皮内注射0．5％罗哌卡因后，主要药动学参数：L1/2α（0．13±0．01）h，T1/2β（1．20±0．08）h，Cmax（4960±13）ng·mL^-1，AUC0→9（11．3±1．6）mg·h·L^-1，AUC0→∞（13．1±2．3）mg·h·L^-1，MRT（1．74±0．13）h。结论建立的LC-MS／MS法快速、准确、灵敏，可用于罗哌卡因豚鼠浸润麻醉的药动学研究。

OBJECTIVE To develop a sensitive and selective LC - MS/MS method for the determination of ropivacaine and pharmacokinetics in guinea pig plasma. METHODS After a simple sample preparation procedure by one - step protein precipitation with acetonitrile,the HPLC separation of ropivacaine and the internal standard bupivacaine was performed on a Zorbax SB -C18 （3.0 150mm,3.5 m） column. The mobile phase consisted of methanol - water （80：20, v/v, containing 5mM ammonium formate）. Detection was performed on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring （MRM） mode via eletrospray ionization （ESI） source. The chromagram was completed in 4 min at 25 ℃. In positive mode, the m/z of ions selected for quantitation were 275.4/126.2 （ropivacaine） and 452. 2/344. 2 （internal standard, bupivacaine）. In the study periods, a single 0. 5% dose of ropivacaine was administered to each guinea pig. RESULTS The method had a lower limit of quantification of 1.0 ng mL^-1. The calibration curve was demonstrated to be linear over the concentration range of 6.58 - 6 580 ng mL^-1. The relative recoveries were 95.6% - 96. 6% , The inter and intra - day precision （ RSD ） were below 6%. The pharmacokinetic parameters of ropivacaine solution was as follows： t1/2α （0.13 ±0.01） h,T1/2β （1.20±0.08） h,Cmax（4 960±13） ng · mL^-1,AUC0-9（11.3 ±1.6） mg·h· L^-1,AUC0-∞（13.1±2.3） mg· h · L^-1 ,MRT（1. 74±0.13） h. CONCLUSION The LC - MS/MS method is proved to be convenient, sensitive, rapid and suitable for pharmacokinetics study.