Derivatisation of amines for electrochemical detection in HPLC

Three derivatising reagents have been studied as possible pre-column derivatisation agents for aliphatic amines for reversed-phase HPLC with electrochemical detection. O-acetylsalicyloyl chloride, N-succinimidyl-3-(4-hydroxyphenyl)propionate, and methyl p-hydroxybenzimidate.HCl were examined. O-acetylsalicyloyl chloride was shown to be a suitable derivatising reagent. Firstly, pure amine derivatives were prepared using the Schotten-Baumann reaction conditions and the structure of the compounds were confirmed by melting point, IR and NMR spectroscopy, and TLC analysis. Analyses were carried out on an ODS-Hypersil column using as mobile phase 50:50 methanol-0.025 M phosphate buffer pH 8 and the derivatives were detected at a glassy carbon electrode set at +0.9 V vs Ag/AgCl. Results for all amine derivatives have indicated the suitability of LCEC detection method with good linearity, precision and sensitivity. Then the amine derivatisation was applied to in situ determination of amines in dilute aqueous solution to give 100% yield, and a linear response down to 10-6 M concentration level. Amine derivatives obtained from N-succinimidyl-3-(4-hydroxyphenyl)propionate and methyl p-hydroxybenzimidate.HCl can also be detected by LCEC but problems were observed with these reagents.