Background: Levothyroxine sodium is widely prescribed to treat primary hypothyroidism. There is consensus that levothyroxine should be taken in the morning on an empty stomach. A pilot study showed that levothyroxine intake at bedtime significantly decreased thyrotropin levels and increased free thyroxine and total triiodothyronine levels. To date, no large randomized trial investigating the best time of levothyroxine intake, including quality-of-life evaluation, has been performed. Methods: To ascertain if levothyroxine intake at bedtime instead of in the morning improves thyroid hormone levels, a randomized double-blind crossover trial was performed between April 1, 2007, and November 30, 2008, among 105 consecutive patients with primary hypothyroidism at Maasstad Hospital Rotterdam in the Netherlands. Patients were instructed during 6 months to take 1 capsule in the morning and 1 capsule at bedtime (one containing levothyroxine and the other a placebo), with a switch after 3 months. Primary outcome measures were thyroid hormone levels; secondary outcome measures were creatinine and lipid levels, body mass index, heart rate, and quality of life. Results: Ninety patients completed the trial and were available for analysis. Compared with morning intake, direct treatment effects when levothyroxine was taken at bedtime were a decrease in thyrotropin level of 1.25 mIU/L (95% confidence interval [CI], 0.60-1.89 mIU/L; P<.001), an increase in free thyroxine level of 0.07 ng/dL (0.02-0.13 ng/dL; P=.01), and an increase in total triiodothyronine level of 6.5 ng/dL (0.9-12.1 ng/dL; P=.02) (to convert thyrotropin level to micrograms per liter, multiply by 1.0; free thyroxine level to picomoles per liter, multiply by 12.871; and total triiodothyronine level to nanomoles per liter, multiply by 0.0154). Secondary outcomes, including quality-of-life questionnaires (36-Item Short Form Health Survey, Hospital Anxiety and Depression Scale, 20-Item Multidimensional Fatigue Inventory, and a symptoms questionnaire), showed no significant changes between morning vs bedtime intake of levothyroxine. Conclusions: Levothyroxine taken at bedtime significantly improved thyroid hormone levels. Quality-of-life variables and plasma lipid levels showed no significant changes with bedtime vs morning intake. Clinicians should consider prescribing levothyroxine intake at bedtime.