Nanosheets have attracted much interest because of their characteristic properties originating from anisotropic and flexible structures. Inorganic nanosheets are synthesized from precursor layered compounds through exfoliation in a liquid phase. In contrast, a versatile exfoliation approach has not been fully studied for organic layered compounds. Here we report two exfoliation approaches for organic layered compounds. Hydrophilic and hydrophobic polydiacetylene (PDA) nanosheets, around 5 nm in thickness, are obtained through exfoliation of the layered precursor in aqueous and nonpolar organic media, respectively. The intercalation of ions and molecules in the interlayer space facilitates swelling and exfoliation. The resultant PDA nanosheets showed characteristic photochemical properties originating from the flexible structure. The exfoliation approach can be applied to a variety of organic layered compounds for the generation of designed nanosheets.