The Effects of Creatine Monohydrate Supplementation to an Egg-Based Dog Spermatozoa Freezing Extender on the Post-Thaw Quality and Functionality
Abstract
Creatine monohydrate (CM) was evaluated as a supplement to a two-step canine freezing extender to determine whether it could improve post-thaw spermatozoa quality by supporting ATP buffering and reducing oxidative stress. Semen from five dogs was cryopreserved using extenders containing 0, 0.5 or 1 mM CM, and post-thaw motility, viability, morphology, velocity and oxidative stress were assessed over 120 minutes. CM produced consistent, biologically coherent improvements across several parameters, with both doses increasing total and progressive motility and enhancing viability numerically, although these trends did not reach statistical significance. The most robust effect was a significant reduction in oxidative stress, with both CM doses increasing the proportion of spermatozoa with <50% NBT staining. Velocity and morphology remained unchanged. Interpretation of dose-dependent patterns suggests that 0.5 mM may offer the most favourable balance between metabolic support and osmotic stability. Limitations included a small sample size, narrow demographic range and reliance on NBT staining for oxidative-stress assessment. Further research should refine the CM dose-response curve, incorporate mitochondrial and osmolarity profiling, and evaluate fertility outcomes through controlled insemination trials. Overall, CM shows promise as a low-dose metabolic additive capable of improving specific functional attributes of post-thaw canine spermatozoa.
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Evidence Hub reads this as a preprint, done in animal, not in people. Its evidence level is 5 of 5 (Very low · case / narrative / animal / in vitro / in silico). Case reports, narrative reviews, and animal, cell or computer-model studies give the weakest evidence for human health.
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