Background Cortisol has a circadian rhythm with an early morning rise, loss of this rhythm is associated with poor health. Our objective was to test the hypothesis that restoring the early morning cortisol rise will improve fatigue and quality of life (QoL) by comparing twice daily Chronocort with once daily Plenadren in patients with adrenal insufficiency. Methods A randomised, double-blind, double-dummy, cross-over study with no washout in 58 patients (29 in each arm) with primary adrenal insufficiency comparing four weeks' Chronocort, 15 mg at night and 10 mg in the morning, a formulation that restores early morning cortisol levels, with four weeks’ once daily Plenadren 25 mg, which only restores daytime cortisol levels. The primary endpoint was the 07:00 h serum cortisol level and secondary endpoints measures of fatigue and QoL. The trial was registered on ClinicalTrials.gov and EU Clinical Trials Register (NCT NCT05222152; Eudract 2021-000144-21), initiated on 11th January 2022 and completed on 18th October 2023. Findings Patients met the primary endpoint and achieved a physiological early morning serum cortisol, median 417 nmol/L on Chronocort versus 6 nmol/L on Plenadren (P < 0.0001). For secondary outcomes the majority of QoL and fatigue measures showed significant benefits for Chronocort including the disease-specific questionnaire AddiQol (P = 0.02), the fatigue questionnaire PROMIS 7b (P = 0.02), SF-36 physical component score (P = 0.01), and EQ-5D-5L (P = 0.02). The Multidimensional Assessment of Fatigue (MAF) was not significantly different between treatments; however, a pre-specified sensitivity analysis showed that in the first treatment period, Chronocort reduced the MAF Score (P = 0.008), suggesting a carry-over effect from period 1 to 2. A post hoc analysis of immune profile in a subset of 19 patients showed that those on Chronocort had an increase in circulating number or frequency of neutrophils, natural killer and natural killer T cells compared to both baseline glucocorticoid treatment and Plenadren treatment. Interpretation Restoring the early morning cortisol levels with twice daily Chronocort 15 mg at night 10 mg in the morning improved health-related quality of life, fatigue and the immune profile compared with 25 mg daily Plenadren. Funding Neurocrine UK Ltd.