Monitoring of the uptake and metabolism of aminooxy analogues of polyamines in cultured cells by high-performance liquid chromatography

Abstract

A high-performance liquid chromatographic method for the determination of polyamines and their aminooxy analogues is described. Oxime derivatization with a ketone is used to protect the aminooxy group during post-column reaction with o-phthalaldehyde. The amount of the polyamines and of the oximes of their aminooxy analogues can be determined simultaneously in cultured cells and cell culture media. The limit of detection is 20-30 pmol, and the response of the fluorescence detection is linear up to 4 nmol. The separation of the aminooxy analogues from the naturally occurring polyamines can be varied by using different ketones for oxime formation. The method was used to measure the stability of aminooxy analogues of putrescine (1-aminooxy-3-aminopropane) and spermidine [N-(2-aminooxyethyl)-1,4-diaminobutane and 1-aminooxy-3-N-(3-aminopropyl)aminopropane] in cell culture media and the uptake into cultured baby hamster kidney (BHK21/C13) cells.