Background: Salicinoids (a type of phenolic glycoside) are plant secondary metabolites with
chemical structures based on salicyl alcohol conjugated to b-D-glucopyranose, with demonstrated antiherbivore
activity. These compounds have been purified and quantified in a variety of contexts. Validation
of published methods is often incomplete, and there is no broadly-applicable reference procedure.
Objective: To develop and validate a robust, versatile procedure for purification and quantification of
salicinoids in salicaceous plants.
Method: We extracted salicinoids from dried, ground Populus foliage into methanol:water, and purified
them by sequential liquid-liquid extraction, flash chromatography and preparative scale HPLC. To
evaluate potential source material for purification of salicinoids, we quantified salicortin, hydroxycyclohexen-
on-oyl salicortin (HCH-salicortin), and tremulacin in methanolic extracts of Populus tremuloides,
P. fremontii, and P. deltoides using ultra-high performance liquid chromatography (UHPLC)
with diode array (DAD) and negative electrospray ionization single quadrupole mass spectrometry
Results: Recovery efficiencies and purities of salicinoids extracted from Populus ranged from 6-63%
and 58->99%, respectively. Both detectors provided accurate quantification of salicinoids; MS was
100× more sensitive than DAD, permitting detection of plant tissue salicinoid concentrations ≥0.0006%
Conclusion: By consolidating and refining existing methods, we developed a reliable, versatile, and
more environmentally-friendly procedure for purification and quantification of salicinoids.