CYP11B1 encodes 11β-hydroxylase, a mitochondrial cytochrome P450 enzyme essential for adrenal corticosteroid biosynthesis 1. The enzyme catalyzes the hydroxylation of 11-deoxycortisol and 11-deoxycorticosterone at the 11β position to produce cortisol and corticosterone, respectively, but cannot produce aldosterone 1. CYP11B1 uses molecular oxygen and requires electrons from NADPH via a mitochondrial electron transfer system comprising FDXR and FDX1/FDX2 1. The enzyme demonstrates regioselectivity preferences for 11β, then 18, and lastly 19 hydroxylation positions. Gene expression is regulated by ACTH and controlled epigenetically through DNA methylation, with hypomethylation associated with increased expression in adrenal adenomas 2. Mutations in CYP11B1 cause 11β-hydroxylase deficiency, the second most common form of congenital adrenal hyperplasia, characterized by impaired cortisol synthesis, androgen excess, and hypertension in approximately 50% of patients due to mineralocorticoid accumulation 34. Additionally, chimeric CYP11B1/CYP11B2 genes cause familial hyperaldosteronism type I (glucocorticoid-remediable aldosteronism), where aldosterone synthesis becomes ACTH-regulated 5. The high sequence identity between CYP11B1 and CYP11B2 presents diagnostic challenges, making CYP11B1 a target for selective therapeutic inhibition strategies 6.