Quantification of ricinine in rat and human urine: A biomarker for ricin exposure

Ricin is a toxalbumin derived from the castor bean plant, Ricinus communis. Ricinine is an alkaloid (3-cyano-4-methoxy-N-methyl-2-pyridone) that shares a common plant source with ricin, and its presence in urine infers ricin exposure. A new quantification method for ricinine was developed that uses solid-phase extraction to prepare 1-mL urine samples (81% recovery) for a 5-min, isocratic high-performance liquid chromatography method, followed by electrospray ionization tandem mass spectrometry. Protonated molecular ions were selected in the multiple reaction monitoring mode and quantified by isotope dilution with 13C6-labelled ricinine as the internal reference. Urine pools enriched with ricinine at two concentrations were characterized as quality control materials and then used to validate the method. The method limit of quantification was 0.083 ng/mL, even with a confirmation ion of low relative abundance. Ricinine was stable in human urine when heated at 90°C for 1 h, and during storage at 25°C and 5°C for 3 weeks. The method was applied to an animal exposure study, a crude ricin preparation scheme, and a forensic analysis. These studies show that ricinine can be measured in rat urine at least 48 h after exposure. Ricinine is present in crude preparations of ricin, and it can be found in human urine after a lethal exposure to ricin.