We present a method for recovering the intrinsic fluorescence coefficient, defined as the product of the fluorophore absorption coefficient and the fluorescence energy yield, of an optically thick, homogeneous, turbid medium from a surface measurement of fluorescence and from knowledge of medium optical properties. The measured fluorescence signal is related to the intrinsic fluorescence coefficient by an optical property dependent path-length factor. A simple expression was developed for the path-length factor, which characterizes the penetration of excitation light and the escape of fluorescence from the medium. Experiments with fluorescent tissue phantoms demonstrated that intrinsic fluorescence line shape could be recovered and that fluorophore concentration could be estimated within ±15%, over a wide range of optical properties.