The significant increase in the production and variety of nanoparticles (NPs) has led to their release into the environment, especially into the marine environment. Titanium dioxide nanoparticles (TiO2-NPs) are used in different industrial sectors, from the food industry to several consumer and household products. Since the aquatic environment is highly sensitive to contamination by TiO2-NPs, this work aimed to give a preliminary assessment of the contamination of packaged seafood, where the food additive TiO2 (E171) is not to be intentionally added. This allowed providing a chemical characterization and quantification of TiO2-NPs in processed canned fish products belonging to different trophic positions of the pelagic compartment and in canned clam. The new emerging technique called single-particle inductively coupled plasma mass spectrometry (spICP-MS) was applied, which allows the determination of nanoparticle number-based concentration, as well as the dissolved titanium. This study highlights how processed food, where the pigment E171 was not intentionally added, contains TiO2 in its nanoparticle form, as well as dissolved titanium. Processed clam represented the seafood with the highest content of TiO2-NPs. In pelagic fish species, we found progressively higher levels and smaller sizes of TiO2-NPs from smaller to larger fish. Our results highlight the importance of planning the characterization and quantification of TiO2-NPs in food both processed and not, as well as where the pigment E171 is intentionally added and not, as it is not the only source of TiO2-NPs. This result represents a solid step toward being able to estimate the real level of dietary exposure to TiO2-NPs for the general population and the related health risks.