The possibility of a diurnal variation in semen quality was tested in 54 human males attending our infertility clinic. Of the enrolled subjects, 24 were normozoospermic and 30 were suffering from oligo- and/or asthenozoospermia. Seminal fluid was collected by masturbation twice by each subject, once in the morning (7:00-7:30 a.m.) and once in the afternoon (5:00-5:30 p.m.). Abstinence from sexual intercourse for 3-4 days before each of the two collections was requested. Semen parameters were evaluated independently by two biologists before and after pellet swim-up. Beside similar macroscopic parameters, specimens collected in the afternoon showed a higher number (P < 0.01) and concentration (P < 0.01) of spermatozoa. Also, immediately (P < 0.05), and at 1 h (P < 0.02) and 2 h (P < 0.01) after pellet swim-up, the number of spermatozoa with progressive linear motility was higher in the afternoon than in the morning. These data are the first documenting a diurnal rhythm in sperm quality which may contribute to the reported variability in semen parameters, and may prove useful for spontaneous and assisted conceptions.