The preferred orientation and conformation of biologically active [D-Ala(2)]- and inactive [L-Ala(2)]Met-enkephalins have been studied by liquid-crystal NMR spectroscopy employing magic-angle- and near-magic-angle-spinning (MAS/NMAS) two-dimensional methodology. The solvent employed was a CsPFO (cesium perfluorooctanoate) liquid crystal so that less proton signals interfere with the spectrum under the MAS/NMAS conditions. The conformation and orientation of [D-Ala(2)]- and [L-Ala(2)]Met-enkephalins were determined in this anisotropic environment from the analysis of the H-1-H-1 direct couplings obtained under the NMAS condition and of the H-1-H-1 ROE factors obtained under the MAS condition. These data were analyzed by the minimization based on pseudoenergy analysis. The results showed that the preferred orientations and the motional restrictions of the tyrosine residues of Met-enkephalin analogues are perpendicular to each other, although the preferred conformations are extended ones for both compounds. Also it is revealed from the NMR analysis that the tyrosine residue is much embedded in the liquid-crystal aggregates for the inactive analogue in contrast to the active one. The interaction between the membrane and the tyrosine residue is pointed out to relate to the activity of enkephalin.