Peptides play an important role in life process.In this paper,two endogenous neuropeptides including methionine enkephalin and leucine enkephalin were selected as research targets.They were synthesized by Fmoc solid phase peptide synthesis strategy.An HPLC-ESI-MS/MS method was established to investigate the composition of synthetic peptides.The possible structure modifications during peptide synthesis procedures were also identified.The results showed that the main byproducts were incomplete peptides and oxidative peptides.Solutions for high efficient synthesis of methionine enkephalin and leucine encephalin were further proposed.This study provides new perspectives in rapid peptide structure identification and efficient bioactive peptide synthesis.