UHPLC-tandem mass spectrometry investigation of andrographolide sodium bisulphite excretion in rats

Objective:To establish an ultra-high performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS) method for quantification of andrographolide sodium bisulphite(ASB) in rat urine and feces,and investigate excretion of ASB in rats after intravenous administration.Methods:ASB was extracted from biological samples using methanol and separated on a Hypersil Gold C18 column(50 mm× 2.1 mm,1.9 μm)with gradient mobile phase consisting of methanol and water at a flow rate of 0.2 mL·min-1.Dehydroandrographolide(DAG) was adopted as an internal standard.Under negative electrospray ionization mode,reaction monitoring of the precursor-product ion transitions m/z 413.2→287.2 was selected for ASB and m/z 331.2→303.3 for DAG.Urine and feces were collected at different time points after intravenously admonition of with ASB at a single dose of 80 mg·kg-1.Results:The linear calibration curve was obtained in the concentration range of 50-5000 ng·mL-1 with a minimal quantification limit of 50 ng·mL-1.Intra-and inter-day precision was less than 11.2% and 13.3%,respectively.Intra-and inter-day accuracy fell in the ranges of 85.8%-101.4% and 87.9%-97.5%,respectively.The recovery ranged from 96.1% to 98.3%,and the matrix effects were between 96.2% and 98.1%.The cumulative amounts of ASB excreted in urine and feces within 72 h as parent drug were 16.9%±3.9% and 34.1%±18.4% of administered dose,respectively.Conclusion:The method is rapid,specific and accurate,which is suitable for ASB quantification in urine and feces and can be applied to ASB excretion study in rats.