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目的:建立一种简便、灵敏的液相色谱串联质谱(LC-MS/MS)方法,测定Beagle犬血浆中泼尼松的浓度,并运用该方法研究泼尼松控释片在健康Beagle犬体内的药代动力学行为。方法:样品预处理采用沉淀蛋白法,内标物为洛索洛芬;采用Venusil ASB C18(2.1 mm×50 mm,3μm)色谱柱,以甲醇-5 mmol·L~(-1)醋酸铵水溶液为流动相进行梯度洗脱,流速为0.2 mL·min~(-1)。离子源为ESI源,采用负离子检测方式,以多反应离子监测(MRM)扫描方式进行检测,检测离子为m/z 357.1→m/z 327.1(泼尼松,待测物),m/z 245.0→m/z 83.0(洛索洛芬,内标物)。6只Beagle犬餐后0.5 h口服给予泼尼松控释片1片(5 mg),于不同时间点分别采集静脉血,测定口服给予泼尼松控释片后Beagle犬血浆中泼尼松的浓度。结果:犬血浆中泼尼松质量浓度在1.00~50.0μg·L~(-1)范围内线性关系良好,定量下限为1.00μg·L~(-1),日内和日间精密度的RSD≤8.9%,平均提取回收率在76.4%~87.5%之间,内标的提取回收率为90.6%,基质效应在87.1%~95.9%之间,内标基质效应为105.5%;结论:Beagle犬口服泼尼松控释片5 mg后,主要药动学参数t_(1/2)为(1.60±0.52)h,k_e为(0.47±0.15)h~(-1),C_(max)为(22.0±2.80)μg·L~(-1),T_(max)为(5.8±0.3)h,AUC_(0-16h)为(93.5±12.7)μg·h·L~(-1)。本文建立的方法可用于泼尼松在Beagle犬体内药代动力学行为的研究。
OBJECTIVE: To establish a simple and sensitive LC-MS / MS method for the determination of prednisone in Beagle dogs plasma and to study the effect of prednisone controlled release tablets in healthy Beagle dogs Pharmacokinetic behavior. Methods: The pretreatment of samples was precipitation protein method, the internal standard was loxoprofen. The samples were separated on a Venusil ASB C18 (2.1 mm × 50 mm, 3 μm) column with methanol-5 mmol·L -1 ammonium acetate aqueous solution The gradient elution was carried out at a flow rate of 0.2 mL · min ~ (-1). The ion source was used as the ESI source, and negative ions were used to detect the ions in m / z 357.1 → m / z 327.1 (prednisone, analyte) and m / z 245.0 in multi-reactive ion monitoring (MRM) → m / z 83.0 (loxoprofen, internal standard). Six Beagle dogs were orally given prednisone controlled-release tablets (5 mg) orally 0.5 h after meal, and venous blood was collected at different time points. Prednisone was measured in Beagle dogs after oral administration of prednisone controlled-release tablets concentration. Results: The prednisone concentration in dogs was linear in the range of 1.00-50.0 μg · L -1 with the lower limit of quantification being 1.00 μg · L -1. The RSDs of intra- and inter-day precision 8.9%, the average recovery was between 76.4% ~ 87.5%, the internal standard extraction recovery was 90.6%, the matrix effect was 87.1% ~ 95.9%, the internal standard matrix effect was 105.5%; Conclusion: The main pharmacokinetic parameters t_ (1/2) were (1.60 ± 0.52) h, k_e was (0.47 ± 0.15) h ~ (-1) and Cmax was (22.0 ± 2.80) μg · L -1, the T max was 5.8 ± 0.3 h, and the AUC 0-16 h was 93.5 ± 12.7 μg · h · L -1. The established method can be used to study the pharmacokinetics of prednisone in Beagle dogs.