The circadian changes in natural killer (NK) activity of peripheral blood mononuclear cells (PDM) were studied in five clinically healthy, diurnally active, nocturnally resting women. Data on spontaneous NK-cell activity were complemented by data on the chronosusceptibility to in vitro inhibition by 1 X 10(-6) M cortisol and by the rhythmometric evaluation of rectal temperature and plasma cortisol as potential circadian markers. In April-July, 1985, blood was drawn at 4-hr intervals for 24 hr starting at 0800 hr. Cells were immediately separated and assayed for NK activity using K 562 cultured cells as a target and a 4-hr 51Cr-release assay. Circadian variations of the spontaneous NK activity were apparent; the maximum of the activity occurred in the morning or in the early afternoon. In individual subjects, peak-to-through differences were 50% or more of the 24-hr mean. Chronosusceptibility to cortisol (20 hr incubation prior to the cytotoxic assay) was ecphasic with respect to the spontaneous NK-cell activity, with a maximum in the evening or night. Data obtained by immunofluorescence using specific anti-NK cell monoclonal antibodies confirm the occurrence of a higher number of phenotypically identifiable NK effectors in the morning vs. other circadian stages. Our data confirm previous findings and extend their scope to immunopharmacology emphasizing the need for time-qualified investigations on immune coordination in vivo.