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Peptide nucleic acids (PNAs) are neutral DNA mimics that bind to complementary DNA or RNA sequences with high affinity and sequence specificity.PNAs developed in the last decades, are particularly promising clue compounds of nucleic acid mimics, in which the natural sugar-phosphate backbone has been replaced by the N-aminoethylglycine units.γ-Carbon modification of the PNA structure allows monomers, and subsequently oligomers, with improved properties to be obtained.In this study, we report the preparation of enantiopure γ-lysine-modified fluorenylmethoxycarbonyl (Fmoc) PNA monomers and the convenient direct enantiomer separation for Fmoc-protected PNA intermediates and monomers without any derivatization for determination of the optical purity.