Quantification of rice alpha-globulin allergen using liquid chromatography-tandem mass spectrometry combined with cysteine-specific modifier and extended stable isotope-labeled peptide

An absolute quantification of alpha-globulin in rice matrices based on the liquid chromatography-tandem mass spectrometry (LC-MS/MS) utilizing extended stable isotope-labeled (SIL) peptides were studied. Two modified signature peptides, derived from the cysteine-specific modification by IAA during the tryptic digestion process, were identified and selected as the surrogates of alpha-globulin due to the 1.5-4.5-fold enhancement of intensity peak compared with its unmodified form. The digestion efficiency could be improved from 72.3-98.8% to 94.0-105.3%, and matrix interference normalized from -26.6-4.5% to -6.0-5.5% when the extended SIL peptides were used as ISs. The limit of detection (LOD) and limit of quantification (LOQ) for the alpha-globulin were 0.25 and 0.8 mu g/g, respectively. The recoveries of alpha-globulin spiked at three levels were between 82 and 105%. The described method was successfully employed for the determination of alpha-globulin in rice and food product samples. Novelty impact statement Two modified peptides were selected as surrogates of alpha-globulin to absolutely quantify the alpha-globulin allergen in rice based on LC-MS/MS. The extended stable isotope-labeled peptide were used as internal standards to normalize variation from the tryptic digestion and matrix effects. The proposed approach showed excellent sensitivity and accuracy, and successfully applied to determination of the alpha-globulin in vary rice and food products.