Little is known about the distribution of the intercalated molecules within the interstratified layers resulting from the pH-dependent interlayer adsorption of ionizable organic molecules, including antibiotics, within smectite-type clay minerals. Here we employed experimental and simulated X-ray diffraction (XRD) to characterize interstratification (or mixed layering) from the intercalation of oxytetracycline (OTC), a commonly used tetracycline antibiotic, within Na-montmorillonite layers at pHs 4, 5, 6, and 8. Our XRD data reveal that OTC is distributed nonrandomly in the interlayers such that Na- and OTC-saturated interlayers coexist. The profile of the full width at half-maximum intensity (fwhm), monitored as a function of increasing layer-to-layer distance (d001), resulting from an increasing amount of intercalated OTC, reflects such mixed-layer crystals under the acidic pH conditions. A minimum in fwhm occurs at a d spacing of about 1.8 nm, which is to be the optimal d001 for OTC-saturated layers, in agreement with molecular modeling results. Using the coordinates of the thermodynamically favorable configuration of the adsorptives in a model montmorillonite interlayer, we simulated XRD profiles to unravel the different patterns of interstratification from the experimental data. At both pHs 4 and 5, Na- and OTC-interlayers are randomly interstratified, whereas at pH 6, these layers are clustered, as characterized by a segregated interstratification pattern. The theoretical layer stacking sequences of the simulated XRD illustrate, as pH increases, the clustering of similar layer types with the exclusion of OTC intercalation from clay populations enriched in Na. At pH 8, both fwhm and d001 indicate OTC adsorption primarily on external surface sites, not within interlayers. Our findings imply that, in addition to chemical speciation, a pH-dependent formation of montmorillonite crystallites with unexfoliated layers may be responsible both for the decreased OTC intercalation and for the increased binding on external sites, thus resulting in the different patterns of interstratification as a function of increasing pH.