Liquid chromatography chloride attachment negative chemical ionization mass spectrometry of diacylglycerol dinitrophenylurethanes

Abstract

Chloride attachment and electron capture negative chemical ionization mass spectra were determined for the 3,5-dinitrophenyl urethanes of natural diacylglycerols following high-performance liquid chromatography on chiral-phase columns using dichloromethane as a source of chloride. For this purpose a chiral column containing the R(+) naphthylethylamine polymer was eluted with an isocratic mixture of either isooctane-f-butyl methyl ether-isopropanol-acetonitrile 80:10:5:5 (solvent A) containing 1% dichloromethane, or with solvent A (20%) and hexane-dichloromethane-ethanol 40:10:1 (solvent B, 80%). The dinitrophenylurethanes gave electron capture molecular and fragment ions in proportions which varied with the composition of the eluting solvent and the ion source pressure. The chloride attachment fragment ions corresponding to the intact diacylglycerols provided a sensitive method of identification and quantitation of the molecular species of the diacylglycerols under all conditions. The detection of the diacylglycerol dinitrophenylurethanes as the negative chloride attachment fragment ions increased the sensitivity and specificity of analysis of the solutes about 100-fold over positive chemical ionization mass spectrometric detection. The detection limit for the dinitrophenylurethanes of natural diacylglycerols was in the low-nanogram range, with the instrument response being linear over two orders of magnitude. For additional structural characterization the dinitrophenylurethanes were subjected to positive chemical ionization mass spectrometry, which resulted in prominent intensities for the [M - DNPU]+ and [RCO + 74]+ ions, but gave no molecular ions. Quantitative analysis of complex mixtures required calibration of the ionization response.