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A rapid, sensitive and selective liquid chromatography/tandem mass spectrometry method (LC-MS/MS) was developed and validated for simultaneous determination of albiflorin and paeoniflorin in rat plasma using geniposide as an internal standard. Plasma samples were extracted by solid-phase extraction. Chromatographic separation was carried out on a Zorbax SB-C(18) analytical column (150 × 2.1 mm × 5 µm) with 0.1% formic acid-acetonitrile (70:30, v/v) as the mobile phase. Detection was performed by multiple reaction monitoring mode using electrospray ionization in the positive ion mode. The total run time was 3.0 min between injections. The calibration curves were linear over a range of 1-1000 ng/mL for albiflorin and 2-2000 ng/mL for paeoniflorin. The overall precision and accuracy for all concentrations of quality controls and standards were better than 15%. Mean recovery was determined to be 87.7% for albiflorin and 88.8% for paeoniflorin. The validated method was successfully applied to the pharmacokinetic study of albiflorin and paeoniflorin in rat plasma after oral administration of Radix Paeoniae Alba extract and Tang-Min-Ling-Wan. The pharmacokinetic parameters showed that albiflorin and paeoniflorin from Tang-Min-Ling-Wan were absorbed more rapidly with higher concentrations in plasma than that from Radix Paeoniae Alba extract. The results provided a meaningful basis for evaluating the clinical applications of traditional Chinese medicine.