In eukaryotes, it is first acylated on its sn-1 position by an ER- or mitochondrial membrane enzyme, glycerol-3-phosphate O-acyltransferase.

A second acyl group is subsequently installed on the sn-2 position making phosphatidic acids.

Glycerol 3-phosphate (G3P) and dihydroxyacetone phosphate (DHAP) are molecules so small that they can permeate the mitochondrial outer membrane through porins and shuttle between two dehydrogenases.

The Glycerol 3-phosphate shuttle is an emergency back-up system to supply neurons' demand of energy.

[2] G3PP activity has subsequently been suggested to have a regulatory role in central metabolism[2] and stress response.