Formation and Verification of the Structure of the 1-Fluorenylmethyl Chloroformate Derivative of Sulfamethazine

Sulfamethazine (SMZ) is derivatized with 1-fluorenylmethyl chloroformate (FMOC) to form the fluorescent adduct SMZ-FMOC. Conditions for formation are optimized with respect to pH, reagent concentration, and reagent ratio. Reagent and product profiles (including the hydrolysis byproduct FMOC-OH) versus time are followed by reversed phase HPLC with UV absorbance detection. FMOC-SMZ has been crystallized, its composition confirmed by microanalysis, and its structure corroborated by IR and NMR spectroscopy. From 10 down to 1 ppm, there is clear gentle curvature in the fluorescence intensity of SMZ-FMOC. The linear response range extends from above 100 ppb down to about 100 ppt, and an increase in sensitivity for the fluorescent detection of FMOC-SMZ (over the usual UV absorbance detection of SMZ) is calculated to be better than 3 orders of magnitude.