In this study, we aimed to investigate lactate excretion in the extracellular conditioned medium of cancer cells treated with anticancer compositions. To explore the missing link in lactylation, we utilized molecular docking and molecular dynamics (MD) simulations to identify potential enzymes capable of generating lactyl-CoA from lactate. Additionally, we conducted molecular interaction studies to determine the binding affinities of lactate and its derived products with HAT p300. Our findings shed light on the significant gaps in our understanding of lactylation, particularly regarding the enzyme that can catalyze lactylation using lactyl-CoA.