PURPOSE To investigate the relationship between intrathyroidal iodine concentration (IC) (mg I/mL) and thyroid hormonal status or pathologic diagnosis with the use of dual-energy computed tomography (DECT). METHODS We retrospectively included patients who underwent neck CT examination between September 2016 and August 2021 using a dual-layer DECT scanner (120 kilovolt peak) for preoperative thyroid imaging. We performed volumetric IC measurements at the thyroid parenchyma on the additional iodine map generated from non-contrast images. We then compared the mean IC of thyroid parenchyma based on thyroid hormonal status (hypothyroid, euthyroid, and hyperthyroid) and diffuse thyroid disease (DTD). Additionally, we determined the accuracy of iodine quantification with our site-specific DECT acquisition protocol using a GammexTM phantom containing seven iodine inserts with different ICs ranging from 2 to 20 mgI/mL. RESULTS Among the 578 patients (M:F: 87:491, age: 48.6 ± 11.7 years) who were finally selected, the mean thyroid parenchymal ICs was the lowest in the hyperthyroid group, followed by the hypothyroid group, and then the euthyroid group (0.68 ± 0.37, n = 44 vs. 1.13 ± 0.42, n = 61 vs. 1.32 ± 0.43, n = 473, P < 0.01, respectively). In the patients with euthyroidism, the mean parenchymal IC was already lower in the patients with pathologically proven DTD than in those without DTD (1.22 ± 0.44 mgI/mL vs. 1.45 ± 0.37 mgI/mL, P < 0.01). Based on the phantom study, the median percentage deviations from the expected values were 5.1% for ICs of 2–20 mgI/mL. CONCLUSION DECT-based IC quantification could be a potentially useful method for identifying patients with thyroid hormone dysfunction or DTD without the use of contrast media. CLINICAL SIGNIFICANCE Without the need for intravenous administration, DECT-based intrathyroidal IC quantification provides potentially valuable information from the non-contrast CT image of the thyroid parenchyma