Simultaneous determination of six alkaloids in blood and urine using a hydrophilic interaction liquid chromatography method coupled with electrospray ionization tandem mass spectrometry

A method was developed for the simultaneous determination of six toxic alkaloids (aconitine, hypaconitine, gelsemine, raceanisodamine, strychnine, brucine) in blood and urine by hydrophilic interaction liquid chromatography (HILIC) coupled with electrospray tandem mass spectrometry. Ephedrine was selected as the internal standard. Samples were extracted and cleaned up by solid-phase extraction (SPE) using Oasis MCX cartridges. Separation parameters such as organic modifier, buffer pH, and concentration of buffer salt were investigated. Gradient separation and analysis were achieved for six alkaloids on a 3-mu m Atlantis HILIC column using a mobile phase consisting of 30 mM ammonium formate and acetonitrile at pH 3. Two multiple reaction monitoring (MRM) transitions for each substance were monitored to provide sufficient identification of alkaloid. The retention mechanisms were explored in the method development. Validation included assessment of linearity, limit of quantification, accuracy, and precision. Bias was less than 15.1% and precision was better than 8.3% for both blood and urine samples. A total of 54 clinical samples were examined by this validated method.