Objective The aim of this study was to determine the extent of drug interactions affecting levothyroxine, using study drugs often co-administered to patients on long-term levothyroxine therapy. Design A retrospective population analysis linking biochemistry and prescription data between 1 January 1993 and 31 December 2012 was used. Patients The study population was Tayside residents prescribed levothyroxine on at least three occasions, within a six month period, prior to the start of a study drug. Individuals acted as their own controls pre- and post-initiation of study drug. Overall 10,999 patients (mean age 58 years, 82% female) being treated with thyroxine were included in the study. Measurements Changes in TSH following initiation of study drug. Results Iron, calcium, proton pump inhibitors, and oestrogen all increased serum TSH concentration: an increase of 0.22 mU/L (p