The biosynthesis of octopamine was determined by the administration of amino acid and amine precursors and enzyme inhibitors and measuring the endogenous levels of octopamine. It appeared to proceed as follows: phenylalanine-tyrosine-tyramine-octopamine. Single and repeated administrations of l-DOPA to rats which were preteated with a monoaminoxidase inhibitor caused a significant increase of octopamine levels in the brain indicating an alternate pathway for octopamine formation from catecholamines. Using dopamine-β-hydroxylase inhibitors it was found that the half life of octopamine in the rat brain is twice as fast as in the heart. α-Methyl-p-tyrosine and 3-iodo-tyrosine, two potent tyrosine hydroxylase inhibitors, were able to reduce the tissue levels of exogenous and endogenous octopamine and noradrenaline.