Tire pyrolysis oil (TPO) is a complex mixture of hydrocarbons, and it is one of the useful fractions obtained from the pyrolysis of waste tires (WT). As a result of its high energy density (HHV ~ 43 MJ/kg), TPO use as a fuel in combustion systems is a promising approach for recycling WT. However, fundamental fuel characteristics and combustion properties of TPO are still unexplored, which stand as a bottleneck for potential applications.This work pursues a comprehensive understanding of the structural characteristics of a TPO produced in a labscale twin-auger reactor as a first step towards defining applications and upgrading strategies. Therefore, advanced analytical techniques such as Fourier Transform -Ion Cyclotron Resonance Mass Spectrometry (FT-ICR MS), and 1 H and 13 C Nuclear Magnetic Resonance (NMR) spectroscopy were utilized. In addition, we also present the characterization of a TPO obtained from adding CaO to WT, as a low-cost catalytic material for its in-situ upgrading, herein named TPO [CaO]. FT-ICR MS results revealed the significant presence of pure hydrocarbons (HC) (HC(TPO) = 74.9 % and HC(TPO[CaO]) = 78.6 %) and hydrocarbons containing one sulfur atom (S1) (S1(TPO) = 14.3 % and S1(TPO[CaO]) = 13.9 %). HC compounds were found mainly in the form of tri-aromatics (26 %), tetra-aromatics (13 and 15 %), and penta-aromatics (22 and 30 %), while S1 compounds in the form of dibenzothiophene (31 %) and benzonaphthothiophene (34 %). The resolved compounds by means of FT-ICR 2 MS exhibited an average double bond equivalent (DBE) number of 11.3 and 12.2 for TPO and TPO[CaO], respectively. These high DBE values were indicators of the significant presence of condensed aromatic structures. 1 H NMR analysis showed that hydrogen atoms in methylene (CH2), methyl (CH3), and naphthenic groups, as well as hydrogen atoms in aromatic structures make up more than 80 % of both fuels. Similarly, carbon atoms in paraffinic groups (both CH2 and CH3) and protonated carbons in aromatic rings together form more than 50 % of the carbon atoms in TPO and TPO [CaO]. The information reported in this work provides new insights into the structural characteristics of the TPO obtained in promising technology as the twin-auger reactor for its use as a fuel, as well as for the design of upgrading strategies.