A rapid, accurate and robust method was firstly developed using ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) assay to quantify kukoamine B, which is a novel drug under clinical development for the treatment of sepsis, in human plasma. Solid-phase extraction (SPE) was used to extract kukoamine B from human plasma. The extracts were separated on a Waters Acquity HSS T3 column (2.1×50mm i.d., 1.8μm) with a gradient elution method, using mobile phases of A (formic acid-water (1:1000, v/v)) and B(formic acid-methanol (1:1000, v/v)). Kukoamine B and internal standard (5-deuterated isotope kukoamine B) were detected under the multiple-reaction monitoring mode by an API 5500 triple quadrupole mass spectrometer with electrospray ionization. The method showed good linearity from 0.100 to 50.0ng/mL according to 1/x weighted linear regression analysis. Inter- and intra-batch precision of kukoamine B were less than 15% and the accuracy was within 85-115%. The extraction recoveries and matrix effect of kukoamine B at three concentration levels were consistent. The sensitivity, specificity and stabilities under various conditions were validated. In conclusion, the validation results showed that this method was rapid, accurate, robust and can successfully fulfill the requirement of clinical pharmacokinetic study of kukoamine B mesylate in Chinese healthy subjects.
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