An easy analytical method for the determination of L-Canavanine in legumes and pulse-based fish feed

L-Canavanine is a naturally occurring non-protein amino acid found in certain legumes, where it plays a significant role in plant defence acting as an anti-herbivory compound by disrupting the biology of insect herbivores and pulse-based fish feed. In this study, a simple, easy and robust analytical method was developed and validated to monitor the compound accurately and reliable. The developed method includes a simple quick and low-cost extraction procedure using acidified acetonitrile. Chromatographic determination is based on Hydrophilic Interaction Liquid Chromatography coupled to tandem mass spectrometry operating in positive electron spray ionization. The method was successfully validated by assessing accuracy, precision, linearity of response (gradient of the calibration curve) at a range of concentrations 1.25 - 25 μg L⁻¹, matrix effect and limit of quantitation according to European Guidelines. One way analysis of variance (ANOVA) was employed for estimating the contribution to overall uncertainty and finally to the reported value/concentration, of homogenization and sub-sampling step.