Vle CONG. INTERN. REPROD. ANIM. INSEM. ARTIF., PARIS, 1968, VOL. II PROTECTIVE EFFECTS OF POLYOLS AGAINST FREEZING INJURY OF BULL SPERMATOZOA II. POLYOLS EFFECTS IN DIFFERENT FREEZING METHODS H. NAGASE AND T, TOMIZUKA National Institute of Animal Industry, Chiba-shi, Japan In the last report (1), the author demonstrated that xylitol gave good protection on survival of bull spermatozoa using isotonic citrate media as a basic solution and the pellet freezing technique. This investigation were made to test the other polyols effects when isotonic citrate solution was used as a basic diluter and frozen in different methods. MATERIALS AND METHODS. Holsteln semen was extended 1:2 with yolk (20%) - 3% citrate media containing various levels of polyols. Freezing was in pellet form, 0.1 ml in size, after cooling to 5°C and equilibrating for 10 hrs. The next trital was conducted to comparing polyols effects in three freezings,slow, pellet and rapid (dropping into liquid nitrogen)., Freezing rates of the semen in the slow (using 0.1 ml of semen in small test tubes) and the pellet were 5-7°C and 25-30°C per minute, respectively. In the rapid freezing, the semen of 0.013-0.015 ml was dropped directly onto liquid nitrogen, and the semen floated on the liquid nitrogen for 20-30 seconds. The semen was extended 1:2 with 0.1M (adnitol, xylitol, sorbitol), 0.5M (all substances tested), 1,0M (erythritol, glycerol, ethylene glycol) and 0.31M (only inositol) which were added to egg-yolk citrate solution and frozen after cooling to 5°C and equilibration for 10 hrs. Motility estimations were made on thawed samples with and without the thawing media (2). RESULTS AND DISCUSSION. Figure 1 represents the post