Neutron diffraction coupled with hydrogen/deuterium isotopic substitution has been used to investigate the structure of a concentrated glycerol water (4:1 mole fraction) solution. The neutron diffraction data were used to constrain a three-dimensional computational model that is experimentally relevant using the empirical potential structure refinement technique. From interrogation of this model, we find that glycerol-glycerol hydrogen bonding is largely unperturbed by the presence of water in the solution. We find that glycerol-water hydrogen bonding is prevalent, suggesting that water molecules effectively take the place of glycerol molecules in this concentrated solution In contrast, we find that water-water hydrogen bonding is significantly perturbed. While the first coordination shell of water in the concentrated solution remains similar to that of pure water, water-water hydrogen bonding is greatly diminished beyond the first neighbor distance. Interestingly, the majority of water molecules exist as single monomers in the concentrated glycerol solution. The preference of isolated water molecules results in a solution that is well mixed with optimal glycerol-water hydrogen bonding. These results highlight the importance of preferential hydrogen bonding in aqueous solutions and suggest a mechanism for cryoprotection by which glycerol effectively hydrogen bonds with water, resulting in a disrupted hydrogen-bonded water network.