Detection of 10 Bufotoxins in Biological Fluids Using UPLC-MS/MS
A method was developed for the detection of 10 bufotoxins(arenobufagin,gamabufotalin,resibufagin,desacetylcinobufagin,resibufogenin,cinobufotalin,bufotalin,cinobufagin,bufalin,and cinobufaginol)in human blood and urine by ultra-high performance liquid chromatography-quadrupole/electrostatic field orbital trap high resolution mass spectrometry(UPLC-Q/Exactive MS).0.8 mL of acetonitrile was added in 0.2mL of blood or urine to precipitate proteins,and then was purified with a Hybird solid phase column.A ACQUITY UPLC HSS T3 column(2.1 mm×100 mm×1.8 µm)was selected.The mobile phase A consisted of 0.1%formic acid in water and B consisted of acetonitrile with a gradient elution program for separation.The mass spectrum was analyzed by electrospray ion source and positive ion mode.The detection limit of 10 bufotoxins in blood and urine can be less than 10ng/mL,and the lowest can reach 0.1 ng/mL.The calibration curves of 10 bufotoxins were in good linear over the range from 20ng/mL to 400ng/mL(R2>0.995)in spiked blood and urine matrix.The extraction recoveries of 10 bufotoxins from blood and urine ranged from 65%to 97.7%,the matrix effect ranged from 90.8%to 109.0%,the intraday precision was within 9.6%,and the interday precision was within 14.7%.The stabilities of 10 bufotoxins in the matrix were good under repeated freezing and thawing condition,room temperature condition,and extraction processing,with the RSD of concentration all within 15%.The method was used to analyze the body fluids collected in animal experiments.The results showed that all 10 bufotoxins had been detected.This method has high sensitivity and broad applicability,and can be used for the identification in bufotoxin poisoning cases.
万方数据
维普
