Complex formation between diheptanoylphosphatidylcholine (DHPC) and beta-cyclodextrin (beta-CD) in deuterium oxide solution has been investigated by measurements of proton NMR chemical shifts and the ROESY spectrum and molecular mechanics calculations. The vicinal coupling constants for protons of alpha-, beta-, and gamma-CDs in their DHPC complexes have been also determined for comparison among these CDs. From the variations in chemical shifts of DHPC and,beta-CD with the addition of beta-CD, the macroscopic equilibrium constant, K-1, of the 1:1 complexation of DHPC and beta-CD is estimated. The magnitude of K-1 is in the order of beta-CD > gamma-CD > alpha-CD for DHPC, although it is in the order of alpha-CD greater than or equal to beta-CD > gamma-CD for single-chain surfactants. From the vicinal coupling constants of the glycerol C1H(2)-C2Hprotons of DHPC, the populations of three rotamers, gauche(+) conformer (G(+)), gauche(-) conformer (G(-)), and trans conformer (T), are estimated. The addition of beta-CD causes a decrease in the T conformer and an increase in the G(+) conformer. These changes are the same as those of gamma-CD but opposite to those of alpha-CD. The microscopic binding constants of CD complexation for the three rotamers are also estimated from the concentration dependence of vicinal coupling constants. The microscopic binding constant for the T conformer is in the order of alpha-CD > beta-CD > gamma-CD, as expected. The presence of DHPC causes large changes in vicinal coupling constants of the,beta-CD protons, suggesting that the P-CD macrocycle is deformed by incorporation of DHPC. A three-dimensional structure of the major complex,beta-CD-G(+) is proposed on the basis of these variations in chemical shift and vicinal coupling constant, the ROESY spectrum, and molecular mechanics calculations: two heptanoyl chains of DHPC are simultaneously incorporated into a beta-CD cavity so that the rim of beta-CD is deformed to an ellipse. This tight contact between the hydrophobic groups of DHPC and beta-CD leads to a large binding constant.