Ethyl propiolate as a post-column derivatization reagent for thiols: development of a green liquid chromatographic method for the determination of glutathione in vegetables.

The present study reports the development, validation and application of a new green liquid chromatographic method for the determination of glutathione (GSH) in vegetable samples. In this work we introduce-for the first time-ethyl propiolate (EP) as an advantageous post-column derivatization reagent for thiolic compounds. GSH (t(R)=6.60 min) and N-acetylcysteine (NAC, internal standard) (t(R)=11.80 min) were separated efficiently from matrix endogenous compounds by using a 100% aqueous mobile phase (0.1%, v/v CH(3)COOH in 1 mmol L(-1) EDTA, Q(V)=0.5 mL min(-1)) and a Prevail(®) reversed phase column that offers the advantage of stable packing material in aqueous mobile phases. The parameters of the post-column reaction (pH, amount concentration of the reagent, flow rates, length of the reaction coil and temperature) were studied. The linear determination range for GSH was 1-200 μmol L(-1) and the LOD was 0.1 μmol L(-1) (S/N=3). Total endogenous GSH was determined in broccoli, potato, asparagus and Brussels sprouts using the standards addition approach. The accuracy was evaluated by both recovery experiments (R=91-110%) and comparison to an o-phthalaldehyde/glycine corroborative post-column derivatization fluorimetric method.