Membrane proteins play crucial roles in cell signaling and transport and, thus, are the targets of many small molecule drugs. The characterization of membrane protein structures poses challenges for the high-resolution biophysical tools because the transmembrane (TM) domain is hydrophobic, opening an opportunity for mass spectrometry (MS)-based footprinting. The hydrophobic reagent diethylpyrocarbonate (DEPC), a heavily studied footprinter for water-soluble proteins, can label up to 30% of surface residues via a straightforward protocol, streamlining the MS-based footprinting workflow. To test its applicability to membrane proteins, we footprinted vitamin K epoxide reductase (VKOR) membrane protein with DEPC. The results demonstrate that besides labeling the hydrophilic extracellular (extramembrane (EM)) domain, DEPC can also diffuse into the hydrophobic TM domain and subsequently label that region. The labeling process was facilitated by tip sonication to enhance reagent diffusion into micelles. We then analyzed the correlation between the residue modification extent and the theoretical accessible surface area percentage (%ASA); the data generally show good correlation with the residue location. Compared with conventional hydrophilic footprinters, the relatively hydrophobic DEPC can map a membrane protein's TM domain, suggesting that the reagent's hydrophobicity can be exploited to obtain structural information on the membrane-spanning region. This encouraging result should assist in the development of more efficient footprinters for membrane protein TM domain footprinting, enabled by further understanding the relationship between a reagent's hydrophobicity and its preferred labeling sites.