Mechanically interlocked polymers can possess significant additional physical properties, in comparison to those associated with their constituent parts. Their unique properties make them attractive for a range of potential applications, such as as biomaterials and molecular machines. Their efficient and reproducible synthesis is therefore of much interest. Both their synthesis and subsequent characterization are intriguing yet demanding. The properties of mechanically interlocked polymeric systems depend not only on the properties of their individual components but also on the topology of the subsequent product. Here traveling wave ion mobility mass spectrometry has been used to investigate the structural properties of a polyrotaxane system. Ion mobility studies reveal that this system remains linear in form with increase in size. Both ion mobility studies and tandem mass spectrometry studies indicate that the macrocycle preferentially remains associated with the ammonium moiety of the polymeric repeat unit and is impeded from moving freely along the axle. This is consistent with NMR observations of the average structure. Analysis of mechanically interlocked polymers by ion mobility mass spectrometry provides additional structural insights into these systems relating to dynamics, heterogeneity, and topology. This molecule-specific information is vital in order to understand the origin of a system's functional properties.