Calculated optimal cooling rates for ram and human sperm cryopreservation fail to conform with empirical observations

The permeability coefficient to water (Lp) and its associated activation energy (Ea) were measured for ram (8.47 microns/min/atm at 25 degrees C, 1.06 kcal/mol) and human (2.89 microns/min/atm at 30 degrees C, 1.93 kcal/mol) spermatozoa. By use of these figures, predictive water loss curves were calculated, from published equations, for different cooling rates from 100 degrees C/min to 100,000 degrees C/min. The calculated curves show that ram spermatozoa cooled at even the fastest rate would be in osmotic equilibrium by -20 degrees C, and human spermatozoa cooled at rates up to 10,000 degrees C/min would be in equilibrium by -15 degrees C. If the nucleation temperature for spermatozoa is taken to be between -20 degrees C and -30 degrees C, then ram and human spermatozoa cooled at these rates would apparently not exhibit any intracellular freezing. There is a significant discrepancy between these calculated optimal cooling rates and the published empirically derived optimal rates of 50 degrees C/min for ram and 10 degrees C/min for human. The failure of ram and human spermatozoa to conform with the established and previously successful model for prediction of optimal cooling rates suggests that damage sustained at high cooling rates may be unrelated to intracellular ice formation.