Guest-host recognition at self-assembled surfaces of two different synthetic calix[4]resorcinarene receptors, a tetrasulfide (R4SC7) and a tetrathiol (R4SH) host, was performed. The host structure was found to impact the molecular organization within the monolayers as well as the interfacial properties. Comparison of structural characterization of the SAMs formed from R4SC7 and R4SH on gold substrates revealed a larger area per molecule for the tetrasulfide host, which indicated that the dialkyl sulfide chains do not pack commensurately below the calix [4] resorcinarene headgroup and can lead to interstitial cavities. As a consequence, the self-assembled surfaces formed from R4SC7 are slightly more hydrophobic and thinner than the SAMs formed from R4SH. Guest-host recognition of a small neutral molecule, alpha-hydroxy-gamma-butyrolactone (HBL), occurred from aqueous solutions on both surfaces on a time scale of several hundred seconds. Adsorption isotherms for the two surfaces revealed that the difference in the free energy change due to adsorption is comparable to thermal energy. Whereas HBL bound to the SAMs formed from R4SH was found to almost completely desorb into an organic solvent, the binding to the R4SC7 surface was largely irreversible. Despite R4SH and R4SC7 possessing macrocyclic cavities of geometrically similar size, the significant difference in the desorption behavior indicates the role of incommensurate packing of the dialkyl sulfide chains below the calix [4] resorcinarene headgroup and the impact of monolayer structure on guest-host recognition at self-assembled surfaces.