This article describes two mathematical formalisms for the determination of the second and fourth order parameters of molecular films using optical spectroscopy. Method A uses polarized total internal reflection fluorescence (TIRF) to calculate the second and fourth order parameters, 〈P 2 (cos θ)〉 and 〈P 4 (cos θ)〉, using an independently determined value for the angle between the absorption and emission dipoles, γ. Method B uses 〈P 2 (cos θ)〉 obtained from attenuated total reflectance (ATR) data, along with polarized TIRF measurements to calculate 〈P 4 (cos θ)〉 and 〈cos 2 γ〉. The choice of a specific method should rely on experimental considerations. We also present a method to separate the contributions of substrate surface roughness and dipole orientation with respect to the molecular axis from the spectroscopically determined second and fourth order parameters. Finally, a maximum entropy approach for construction of an orientation distribution from order parameters is compared with the commonly used delta and Gaussian distributions.