3H‐leucine incorporation was measured in a cell‐free system from rat liver. Even under conditions of constant amino acid concentrations and energy supply, we observed a diurnal rhythmicity when analysing tissue samples from serially sacrificied animals. Maximum incorporation rates were found during dark‐time, whereas a characteristic minimum occurred in the second half of light‐time. Injections of glucagon 20 min prior to killing significantly increased the incorporation rates. The degree of stimulation depended on the phase of the diurnal cycle; the periodicity of stimulation by glucagon approximately paralleled the rhythm of unstimulated incorporation. Despite of the stimulatory effect of glucagon administrationinvivo,addition of cAMP (and also cGMP) to thein vitrosystem resulted in an inhibition, which represents, however, a well‐known unphysiologic phenomenon. Nevertheless, the extent of inhibition also varied within the diurnal cycle; these changes were due to differing temporal inhibition kinetics.