Determination of Double Bond Positions in Unsaturated Fatty Acids using Permanganate Oxidation and Tandem Mass Spectrometry of CuIICl+ Adducted Ions

Positional characterization of C═C bonds is crucial for elucidating the structure of unsaturated fatty acids (UFAs). Chemical derivatization of C═C bonds prior to tandem mass spectrometry (MS/MS) has proven to be an effective approach for producing diagnostic fragments that enable precise determination of C═C locations. This study employed neutral potassium permanganate to convert C═C bonds of unsaturated fatty acids into α-hydroxy ketones, which were subsequently analyzed by low-energy collision-induced dissociation (CID). Negative-ion mode CID of deprotonated mono-oxidized unsaturated fatty acids generated diagnostic fragment ions alongside abundant false-positive peaks. Various metal ions (including alkali and transition metals) were then evaluated as charge carriers for mono-oxidized unsaturated fatty acids. Notably, positive-ion mode CID of CuIICl+ adducted ions, [(M+2O)+CuIICl]+, produced abundant diagnostic fragments for identifying C═C bond positions in all tested fatty acids with varying degrees of unsaturation (conjugated/unconjugated C═C bonds), without any interference from false-positive fragments. This method was also successfully applied to the analysis of total UFAs in human plasma, providing information on the number and position(s) of carbon-carbon double bond(s).