Objective: To evaluate cryodamage on human sperm motility, cryosurvival rate, and sperm chromatin assessed by acridine orange staining method (AO test) after a six-month freeze-thawing process using liquid nitrogen vapor.

Study design: Experimental study.

Material and Method: Twenty normal semen samples were obtained from the male partner of infertile couples attending the infertility unit, Siriraj Hospital. After semen analysis, each semen sample was frozen with liquid nitrogen vapor. The acridine orange test was used for assessment of chromatin structures. After 6 months of cryostorage, semen samples were thawed and the effects of cryopreservation on sperm chromatin integrity, motility, morphology, vitality, and cryosurvival rate were evaluated.

Results: The mean percentage of normally condensed sperm chromatin in the native semen sample decreased significantly (87.3±9.1 vs 47.9±26.2; p<0.001) after the freeze-thawing process using liquid nitrogen vapor. Furthermore, the mean percentage of sperm motility and vitality also decreased significantly after the freeze-thawing process (52.6±1.9 vs 23.2±10.6 and 78.7±5.6 vs 30.3±8.8 respectively; p<0.001). In contrast, the numbers of sperm with normal morphology after cryopreservation were not different from those before the procedure (21.4±4.3 vs 24.2±23.9; p = 0.606).

Conclusion: The freeze-thawing procedure using liquid nitrogen vapor had effects on chromatin, motility, and vitality of human spermatozoa. The six-month cryopreservation of semen is a good method for avoiding the window period of HIV; however, this can cause a lot of damage to spermatozoa, thus, limits their further use in the treatment of infertility.

Keywords: Liquid nitrogen vapour, Sperm DNA integrity, Cryopreservation, AO test