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Post‐ejaculatory increase in human semen osmolality in vitro

Spermatozoa prepared in vitro for assisted reproductive technology (ART) encounter other challenges than in vivo. One could be to survive and function despite varying extracellular osmolality. There is a lack of consistent knowledge of human semen osmolality and changes after liquefaction. The aim of this study was to investigate changes in semen osmolality that may occur in vitro after ejaculation and during sperm handling for ART. Osmolality was measured in 86 semen samples by freezing‐point depression during storage in vitro at various times and temperatures. The freezing‐point depression method was robust and reliable for measuring the osmolality of whole semen as shown by a low variability between repeats. At ejaculation, the osmolality was isotonic to cervical mucus and body fluids. There was a marked increase in osmolality after liquefaction, and the degree of increase varied greatly between samples. Osmolality rose with increasing temperature, and the progressive increase was blocked by denaturising temperature. Spermatozoa in each individual semen sample experienced a highly variable environment in vitro with respect to osmolality. This may be of importance regarding the handling of semen samples for the outcome of ART.