Washed spermatozoa from 8 semen samples were used and
incubated in the described solutions for 72 h at 4 °C and at a cell
density of 5–7× 107 cells ml−1
. Immediately after dilution and after
72 h incubation, subsamples were taken and ATP was extracted with
3 mol/l perchloric acid. Using the UV-spectrophotometric assay of
Bergmeyer (1985) the samples were centrifuged to remove precipitated
protein, neutralized and ATP levels were analyzed