This article reports a novel method to tune the photo-luminance properties of ammonium 1-anilinonaphthalene-8-sulfonate (ANS) in a 2D matrix of layered double hydroxide (LDH) by changing the interlayer microenvironment. ANS and a series of surfactants with different alkyl chain lengths (pentanesulphonate (PES), hexanesulphonate (HES), heptanesulphonate (HPS), decanesulphonate (DES), and dodecylsulphonate (DDS)) were respectively cointercalated into the galleries of ZnAl-LDH by the anion exchange method. Thin films of ANS/surfactant-LDHs obtained by the solvent evaporation method possess good c orientation as revealed by XRD and SEM. It was found that the ANS/HPS-LDH film showed the maximum fluorescence efficiency and the longest intensity-average lifetime among these ANS/surfactant-LDH composites owing to the "size-matching" rule between the organic dye and surfactant. Moreover, the fluorescence properties can be tuned by changing the relative molar ratio of ANS/HPS, and the film containing 20% ANS (molar percentage, expressed as ANS(20%)/HPS-LDH) exhibits the maximum fluorescence efficiency, the longest average lifetime, and significantly enhanced photo and thermal stability. In addition, the composite films show fluorescence anisotropy, attributed to the preferential orientation of ANS in the LDH gallery. Therefore, this work demonstrates a feasible approach to tuning the photoluminescence properties of a dye confined in an inorganic 2D matrix via changing the interlayer microenvironment, which may be considered to be a good candidate for solid photoluminescence materials, nonlinear optics, and polarized luminescence materials.