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Most biomolecules are few non-fluorescent. Therefore, highperformance liquid chromatography (HPLC)-fluorescence detection is a powerful tool for highly sensitive and reproducible quantification analysis. Recently, we used a water-soluble, thiol-specific fluorogenic reagent (ammonium 7-fluoro-2,1,3-benzoxadiazole-4sulfonate (SBD-F) or 7-chloro-N-[2-(dimethylamino)ethyl]-2,1,3benzoxadiazole-4-sulfonamide (DAABD-Cl)) for the derivatization of proteins. These reagents have no fluorescence themselves but produce fluorescent proteins when derivatized (FD), resulting in a low background noise and thus high signal-to-noise ratio in the detection of the derivatized proteins. DAABD-Cl is a rather small molecule with an ionizable group in the neutral medium at pH 7 and is highly reactive to the thiol moiety of cysteine in proteins [1]. Additionally, the detectability of DAABD derivatives is significantly enhanced, compared with SBD derivatives in MS/MS analysis, probably due to the positively charged group in the DAABD moiety.

Development of Highly Sensitive and Reproducible Analytical System for Biological Macromolecules with Fluorogenic Derivatization