We report on the use of surface second harmonic generation (SHG) measurements to probe microscopic organization in asymmetrically bound aliphatic multilayer assemblies. The use of layers that are bound asymmetrically using metal ion coordination chemistry allows the orientation of the individual layer constituents to be controlled, and the asymmetric coordination makes the assemblies chi ((2))-active. We find that, for the systems we report here, the surface second-order nonlinear response increases in proportion to the square of the number of layers initially and then saturates and decreases with additional adlayers. Optical null ellipsometry and FTIR data point to regular growth in terms of thickness and chromophore density, and the surface SHG data point to increasing disorder with the adsorption of additional layers. The FTIR data suggest a subtle decrease in overall order of these chains with increasing number of layers, but this trend is not as pronounced as that seen in the surface SHG data. We interpret these data in the context of an increase in the distribution of nonlinear chromophore orientations with layer growth and not a change in the average orientation angle.