When used for automotive antifreeze, glycerol has the advantage of being non-toxic, withstanding relatively high temperatures and being corrosive.

Like ethylene glycol and propylene glycol, glycerol is a non-ionic cosmotrope that competes with water-molecule hydrogen bonds, forming strong hydrogen bonds with water molecules. This disrupts the crystal lattice formation of the ice unless the temperature drops significantly. The minimum freezing point temperature is approximately −36 ° F / −37.8 ° C, which corresponds to 60-70% glycerol in water. It was used as an antifreeze for automotive applications before being replaced with ethylene glycol, which has a lower freezing point. Although the minimum freezing point of a glycerol-water mixture is higher than an ethylene glycol-water mixture, glycerol is non-toxic and is being re-examined for use in automotive applications. It is designed for use as antifreeze in many sprinkler systems.

In the laboratory, glycerol is a common component of solvents used for enzymatic reagents stored at temperatures below 0 ° C depending on the freezing temperature of solutions containing high concentrations of glycerol. It is also used as a cryoprotectant where glycerol dissolves in water to reduce the damage caused by ice crystals to laboratory organisms stored in frozen solutions such as bacteria, nematodes and mammalian embryos.