Because acids in petroleum materials are known to corrode processing equipment, highly acidic oils are sold at a discount [on the basis of their total acid number (TAN)]. Here, we identify the acidic species in raw Canadian bitumen (Athabasca oil sands) and its distilled heavy vacuum gas oil (HVGO) as well as acid-only and acid-free fractions isolated by use of an ion-exchange resin (acid-IER) and negative-ion electrospray ionization Fourier transform ion cyclotron resonance (ESI FT-ICR MS) mass spectrometry. The ultrahigh mass resolving power (m/Delta m(50%) > 400 000) and high mass accuracy (better than 500 ppb) of FT-ICR MS, along with Kendrick mass sorting, enable the assignment of a unique elemental composition to each peak in the mass spectrum. Acidic species are characterized by class (NnOoSs heteroatom content), type [number of rings plus double bonds to carbon or double-bond equivalent (DBE)], and carbon number distribution. We conclude that the analytical capability of FT-ICR MS and the selectivity of the ESI process eliminate the need for acid fractionation to characterize naphthenic acids in bitumen. However, because the acid-free fraction (not retained on the acid-IER) contains SxOy heteroatomic classes not observed in the parent bitumen, acid-IER fractionation does help to identify such low-abundance species. Further, we observe that a subset of the acids identified in the parent bitumen distill into the HVGO fraction. Variations in the carbon number and aromaticity of the classes are discussed in detail.