We manipulate the negative fixed charge density Qf at the c-Si/Al2O3 interface by applying a bias voltage in a metal-oxide-semiconductor configuration or by depositing corona charges onto the Al2O3 film. A significant increase of the negative fixed charge density from |Qf| = 4 × 1012 cm−2 to values above 1013 cm−2 is observed for surface Fermi energies close to or within the silicon conduction band. The additional charges are shown to be partly unstable under annealing or changing the polarity of the bias voltage. Our experimental data are best described by assuming at least three different types of charge traps responsible for the formation of the negative fixed charge density at the c-Si/Al2O3 interface.