Simultaneous analysis of 40 pyrrolizidine alkaloids in herbs and spices by supercritical fluid chromatography-tandem mass spectrometry.

Pyrrolizidine alkaloids, including their free bases (PAs) and their N-oxides (PANOs), are plant-derived natural toxins, which often pose analytical challenges because they have many structurally similar isomers that are difficult to separate by conventional chromatography. To address this issue, we employed supercritical fluid chromatography-tandem mass spectrometry. A 3-hydroxyphenyl column with an ammonium formate-containing methanolic modifier gradient enabled simultaneous determination of the 40 PAs/PANOs, although some isomeric pairs were only partially resolved. We validated the developed method using nutmeg as a matrix. The calibration curves showed good linearity over the range of 1 to 100 μg/kg, with coefficients of determination of 0.997-1.000 for all PAs/PANOs. Recoveries were 84.9-101.3%, and intra- and inter-day precisions were 2.0-16.3% and 3.0-16.7%, respectively, at the spiking level of 1 μg/kg. These validation results indicate that the present method can quantify 40 PAs/PANOs in herbs and spices. We also determined PAs/PANOs in commercially available herbs and spices (oregano, savory, thyme, marjoram, cumin, dill seed, and sage) to confirm the applicability of the developed method. At least four PAs/PANOs were quantified in all samples other than dill seed. Relatively high levels of PAs/PANOs (>1,000 μg/kg) were noted in oregano and cumin. These results suggest our developed analytical method is a reliable and useful means for determining PAs/PANOs in herbs and spices.