In New Zealand, 95% of the semen used for artificial insemination in cattle is processed as liquid semen. Storage of liquid semen for up to 3 days in Caprogen (R) diluent enables a 10-fold reduction of the insemination dose, compared to frozen-thawed semen, without a reduction in fertility. In this Caprogen (R) diluent spermatozoa are stored under N-2 gas in the presence of catalase. However, a new diluent (CEP-2), which was originally based on the biochemical composition of bovine cauda epididymal plasma, could become an appropriate alternative to Caprogen (R). In this study, the effect of addition of catalase to bovine spermatozoa stored for 6 days in CEP-2 diluent under aerobic and anaerobic conditions was evaluated and compared with a Tris diluent. Additionally, the quality and in vitro fertilizing capacity of fresh bovine semen stored for 6 days at 5 degrees C in the Trilady (R), CEP-2 (without catalase and N-2 gas) and Caprogen (R) diluent were compared. Addition of 4.5 mg/mL catalase to CEP-2 diluent under aerobic and anaerobic conditions had no effect on sperm quality. Spermatozoa stored in CEP-2 diluent moved faster and straighter than spermatozoa stored in Triladyl (R) or Caprogen (R) diluent. The in vitro fertilization and polyspermy rates did not differ significantly between spermatozoa stored for 6 days at 5 T in CEP-2 and Caprogen (R) diluent, but were significantly lower for spermatozoa stored in Triladyl (R) diluent. We can conclude that based on the in vitro results, the CEP-2 diluent is a better diluent than Triladyl (R) and a good alternative to the Caprogen (R) diluent for long term storage of fresh bovine semen at 5 T. To confirm these promising in vitro results further in vivo experiments are required. (c) 2004 Elsevier Inc. All rights reserved.