Protective Effects of Serotonin, L-Carnitine, and Coenzyme Q10 on Human Sperm Cryopreservation: Functional and In Silico Evidence

Background: Sperm cryopreservation can impair sperm function through oxidative stress, membrane damage, mitochondrial dysfunction, and DNA fragmentation. This study evaluated the effects of supplementing a basal freezing medium with serotonin (5-HT), L-carnitine (L-C), and coenzyme Q10 (CoQ10) on post-thaw human sperm quality and explored potential molecular interactions using molecular docking. Methods: Semen samples from 63 normozoospermic volunteers were processed using the swim-up technique, and each sample was divided into five aliquots assigned to the following conditions: commercial freezing medium; basal freezing medium; basal medium supplemented with 5-HT alone; basal medium supplemented with L-C and CoQ10; and basal medium supplemented with all three compounds. Post-thaw motility, intracellular ROS, viability, and apoptosis-associated changes were assessed in all 63 donors; DNA fragmentation was assessed in a subset of 32 donors. Molecular docking explored predicted interactions of 5-HT, L-C, and coenzyme Q1 (CoQ1), a short-chain CoQ10 analogue, with selected sperm-related targets. Results: The combined formulation showed the highest median total (54%) and progressive (39%) motility, the lowest median percentage of DCF-positive spermatozoa (11.30%), and the lowest median DNA fragmentation index (2.58%). Pairwise analyses showed significant differences between the combined formulation and several comparator groups, depending on the outcome. Viability was significantly higher with the combined formulation than with basal, 5-HT-supplemented, and L-C/CoQ10-supplemented media (all Bonferroni-adjusted p < 0.001). Docking predicted interactions of 5-HT with HTR4 and SLC6A4 and of CoQ1 with CPT1A. Conclusions: Combined 5-HT, L-C, and CoQ10 supplementation was associated with favorable changes in several post-thaw sperm quality parameters. These findings support further investigation of this formulation, while the predicted molecular interactions require experimental validation.