Nanoflow Gradient Generator Coupled with (mu)-LC-ESI-MS/MS for Protein Identification

Reversed phase high-performance liquid chromatographic (RP-HPLC) methods are proposed for the analysis of phenol, thymol, chlorocresol and chloroxylenol in commercial pharmaceutical dosage forms. The use of 1-fluoro-2,4-dinitrobenzene as UV labelling reagent in pre-column derivatization has been investigated. The compound reacts rapidly (2 min) under mild conditions (ambient temperature or 40°C) with phenols to give ethers that can be separated by RP-HPLC and detected at 292 nm. The other procedure involves a post-column on-line photochemical conversion. A photoreactor was arranged between the analytical column and the fluorescence (lambdaex=270 nm, lambdaem=310 nm) and UV-diode array (lambda=270 nm) detector to enhance the performance of the methods. Additional informations of the analyte structure and photoreactivity by UV spectra (photoreactor `onʹ and `offʹ) were obtained. The methods showed good selectivity and sensitivity.