We present an on-demand electrochemical mass spectrometry workflow for lipid structural characterization with the capability of identifying double-bond and
sn-positional isomers. Two voltage-controlled electrochemical reactions, interfacial electro-epoxidation and anodic corrosion of a cobalt electrode, are cascaded in this strategy. The epoxidized products and Co-adducted ions of lipids are fragmented in tandem MS to generate structure characteristic fragments that indicate double-bond and sn-positions. The unique feature of this workflow lies in the sequential localization of double bonds, elucidation of sn-positions, and collection of native lipid information in a single system and simply by tuning ESI voltages. This feature, as well as low sample consumption, no need for extra apparatus, and quantitative analysis, should allow wide applications in the lipidomic field.