The advantages of using a combination of LDL and glutamine in comparison with TRIS egg yolk and Equex® STAMP extenders in the cryopreservation of canine semen

Twenty semen samples taken from 5 dogs were frozen in liquid nitrogen at −196 °C in four different extenders: one control extender based on 20% egg yolk, 6% LDL alone (low density lipoproteins: the active cryoprotective principle in chicken egg yolk), 6% LDL combined with 20 mmol glutamine, and Equex® (a reference extender that we wish to compare with the LDL-glutamine combination). After thawing, spermatozoal motility was evaluated using a HAMILTON THORNE CERROS 12 image analyzer; the percentage of motile spermatozoa was 27.7% in the egg yolk extender (p < 0.05), 49.9% with 6% LDL alone (p > 0.05), 54.7% in the 6% LDL + 20 mmol glutamine extender, and 47.9% with Equex® (p > 0.05). The motility parameters (VAP, VCL, VSL and ALH) were also superior in the 6% LDL + 20 mmol glutamine extender in comparison with the other extenders.Finally, the spermatozoa were generally better protected during freezing with the 6% LDL + 20 mmol glutamine association than with the egg yolk, 6% LDL, or Equex extenders in terms of the flagellar plasma membrane (HOS test), DNA (Acridine orange test), and acrosome integrity (Spermac® test: no significant difference). The Equex® extender obtained the best results for the acrosome, followed by 6% LDL + 20 mmol glutamine (FITC-PSA test: p < 0.05 between each extender).