Ramesh Bokka, Manjula Nemali, Ramakrishna Sistla, Devi Potturi Sita
Natural Products Chemistry Division, Indian Institute of Chemical Technology, Tarnaka, Hyderabad 500607, India.
Medicinal Chemistry & Pharmacology Division, Indian Institute of Chemical Technology, Tarnaka, Hyderabad 500607, India.
J Pharm Anal. 2015 Feb;5(1):43-50. doi: 10.1016/j.jpha.2014.05.001. Epub 2014 May 20.
A novel bioanalytical method was developed and validated for the quantitative determination of darunavir (DRV) in rat plasma by employing hydrophilic interaction chromatography and tandem mass spectrometry (HILIC-MS/MS) with supported liquid extraction (SLE). Irbesartan (IRB) was used as an internal standard (IS). The analyte in rat plasma (200 µL) was isolated through SLE using ethyl acetate as the eluting solvent. The chromatographic separation was achieved on Luna-HILIC (250 mm×4.6 mm, 5 μm) column with a mobile phase of 0.1% of formic acid in water:acetonitrile (5: 95, v/v), at a constant flow rate of 1.0 mL/min. The MS/MS ion transitions for DRV (548.1→392.0) and IS (429.2→207.1) were monitored on an ion trap mass spectrometer, operating in the multiple reaction monitoring (MRM) mode. The lower limit of quantitation (LLOQ) was 0.2 ng/mL and quantitation range was 0.2-5000 ng/mL. The method was validated for its selectivity, sensitivity, carryover, linearity, precision, accuracy, recovery, matrix effect and stability. The method was successfully applied to pharmacokinetic study in rats.
建立了一种新型生物分析方法，并通过亲水相互作用色谱-串联质谱法（HILIC-MS/MS）结合支持液液萃取（SLE）对大鼠血浆中的达芦那韦（DRV）进行定量测定。厄贝沙坦（IRB）用作内标（IS）。采用乙酸乙酯作为洗脱溶剂，通过SLE从200 μL大鼠血浆中分离出分析物。在Luna-HILIC（250 mm×4.6 mm，5 μm）柱上进行色谱分离，流动相为0.1%甲酸水溶液：乙腈（5:95，v/v），流速为1.0 mL/min恒定。在离子阱质谱仪上以多反应监测（MRM）模式监测DRV（548.1→392.0）和IS（429.2→207.1）的MS/MS离子跃迁。定量下限（LLOQ）为0.2 ng/mL，定量范围为0.2 - 5000 ng/mL。该方法在选择性、灵敏度、残留、线性、精密度、准确度、回收率、基质效应和稳定性方面进行了验证。该方法成功应用于大鼠的药代动力学研究。
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