Hydrophilic interaction chromatography (HILIC) coupled with tandem mass spectrometry was developed for the quantification of swainsonine in plants. Optimized separation was performed on an Acquity UPLC BEH HILIC column using acetonitrile/water (90:10, v/v) with 10 mmol/L formic acid as the mobile phase. In comparison to the conventional reversed-phase separation mode, the HILIC technique provided a significantly better retentivity for swainsonine. The calibration curve showed a good linearity over the concentration range 5–750 μg/L corresponding to 0.75–112.5 μg/g in the dry matter (DM) of plant material. The limit of quantification for the method was 5.0 μg/L (0.75 μg/g DM). The mean recoveries of swainsonine in the two milkwetch matrices ranged from 85.5 to 104.4%. The method was applied to determine swainsonine in two milkvetch species (Astragalus cicer and Astragalus glycyphyllos) growing in Lithuania. None of the samples contained detectable amounts of swainsonine.

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