A simple and mild derivatization method for hydroxyl compounds using condensation agent carbazole-9-N-(2-methyl)-acetyl-benzene-disulfonate and its application for the determination of volatile alcohols from plasma by HPLC with fluorescence detection

摘要

A simple and sensitive HPLC method for the determination of hydroxyl compounds including volatile alcohols from spiked plasma, using a fluorescent condensation agent, namely carbazole-9-N-(2-methyl)-acetyl-benzene-disulfonate (CMABS), has been developed. A mixture of alcohols and pyridine in dichloromethane was treated with CMABS to give a quantitative yield of esters. The maximum fluorescence emissions for the derivatized alcohols are at 365 nm (#lambda#_(ex) 335 nm). Studies on derivatization conditions indicate that alcohols react very fast with CMABS in presence of pyridine in dichloromethane to give the corresponding fluorescent derivatives. The method, in conjunction with a multi-step gradient, offers a baseline resolution of the common alcohol derivatives on a reversed-phase C_(18) column, which is more convenient and more efficient than previous methods which require the prior conversion of the carboxylic acids to the acyl chlorides. The application of this method to the analysis of volatile alcohols in plasma was also investigated. The LC separation shows good selectivity and reproducibility for alcohol derivatives. The relative standard deviations (n = 5) for 100 pmol of each alcohol are <4%. The detection limits are at the fmol level for C_1-C_3 alcohols and pmol level for C_4-C_9 alcohols.