The conformational equilibrium of the general anesthetic propofol (2,6-diisopropylphenol) has been studied in a supersonic expansion using broadband chirped-pulse microwave spectroscopy. Three conformers originated by the combined internal rotation of the hydroxyl and the two isopropyl groups have been detected in the jet-cooled rotational spectrum. The most stable conformer exhibits tunneling splittings associated with the internal rotation of the hydroxyl group, from which we determined the torsional potential and barrier heights (905-940 cm(-1)). The carbon backbone structure was derived from the spectral assignments of all 12 (13)C monosubtituted isotopologues in natural abundance and confirmed a plane-symmetric gauche orientation of the two isopropyl groups (Gg) for this conformer. In the other two detected conformers (EG and GE) one of the isopropyl groups is eclipsed with respect to the ring plane while the other is gauche, differing in a ∼180° rotation of the hydroxyl group. Supporting ab initio calculations provided information on the potential energy surface and molecular properties of the title compound.