A He-Ne laser (632.8 nm), a laser diode (648 nm), and a light emitting diode (678 nm) have been used to obtain anti-Stokes fluorescence of phenalemine 512 in a polymeric matrix (PMMA) with concentrations of 10-3 - 10-2 M. The existence of a "red limit" of excitation of anti-Stokes fluorescence is shown experimentally and theoretically. The temperature dependence demonstrates the decrease in the intensity of anti-Stokes fluorescence with the decrease of temperature. The energy of activation of anti-Stokes fluorescence of phenalemine 512 in PMMA for simple molecule was determined to be (2.52 ± 0.09) 10-20 J.