Polycyclic aromatic sulfur heterocycles are undesirable compounds in fuel and refined petroleum-based products. The sulfur compounds in diesel are limited strictly to an ultralow concentration according to environmental protection rules. Alkyldibenzothiophenes are recalcitrant species in the hydrodesulfurization process, and speciation of dibenzothiophenes in deeply hydrodesulfurized diesel fuel provides important information to the catalyst and process improvement. However, most of the existing analytical methods are unsuitable for identification of ultralow-sulfur compounds in deeply hydrodesulfurized diesel because of the limits of sensitivity. A new method for direct quantitative analysis of alkyldibenzothiophenes in deeply hydrodesulfurized diesel fuel was developed by gas chromatography quadrupole time-of-flight mass spectrometry, which exhibited good linear response and high precision and sensitivity. The practicability of the method was shown by an extensive qualitative and quantitative analysis of alkyldibenzothiophenes in hydrodesulfurized middle distillates at different sulfur levels.