Optical homogeneous linewidths Γ hom of organic molecules in glassy hosts can be determined by means of hole burning. This process may either be the result of photochemistry (photochemical hole burning) or of a relative reorientation of the guest-host system (nonphotochemical hole burning) on laser excitation. The experiments described here have been performed on the 0−0 S 1←S 0 transitions of a large variety of amorphous and semi-crystalline systems at temperatures between 0.3 and 20 K. It was found that, independent of the hole-burning mechanism, Γ hom follows a T 1.3 temperature law, and extrapolates to or in fact attains the fluorescence lifetime-limited value of the guest for T → 0. From a study of the holewidth dependence on laser power, burning time, sample preparation and detection method, conditions could be established for the determination of Γ hom and the study of spectral diffusion. Semi-crystalline materials, as opposed to amorphous systems, have a much steeper Γ hom dependence on temperature than T 1.3 which varies with the degree of crystallinity of the polymer host. The results show that hole burning is a very sensitive technique to probe the amount of disorder of the environment directly surrounding the guest molecule. An interpretation of the experiments in terms of various theoretical models for dephasing in glasses is presented.