The adsorption of hexadecyltrimethylammonium (HDTMA) in smectite was studied by adsorption isotherms, X-ray diffraction (XRD) and high-resolution transmission electron microscopy (HRTEM). Smectites that had reacted for 48 h with HDTMA cations equivalent to 0.2–3.0 times the cation exchange capacity (CEC) were converted to HDTMA-exchanged smectites with various d-spacings. Study of HDTMA-smectites by HRTEM suggests that the HDTMA adsorption results in interlayer expansion with various d-spacings and irregular wavy layer structures. We believe that HDTMA loading beyond the CEC of smectite affects the structure of clay by the additional adsorption of HDTMA-Br− via hydrophobic bonding. Surfactant orientation probably depends on the quantity of surfactant in the interlayer. Our TEM study shows that the structure of the adsorbed HDTMA layer in the interlayers of smectite depends on the charge distribution and chemical composition of smectite.