The main aim of origins of life research is to find a plausible sequence of transitions from prebiotic chemistry to nascent biology. In this context, understanding how and when phospholipid membranes appeared on early Earth is critical to elucidating the prebiotic pathways that led to the emergence of primitive cells. Here we show that exposing glycerol-2-phosphate to acylating agents leads to the formation of a library of acylglycerol-phosphates. Medium-chain acylglycerol-phosphates were found to self-assemble into vesicles stable across a wide range of conditions and capable of retaining mono- and oligonucleotides. Starting with a mixture of activated carboxylic acids of different lengths, iterative cycling of acylation and hydrolysis steps allowed for the selection of longer-chain acylglycerol-phosphates. Our results suggest that a selection pathway based on energy-dissipative cycling could have driven the selective synthesis of phospholipids on early Earth.