Hybrid heterostructures are crucial in photovoltaics where the overall efficiencies of the materials are closely related to the level of charge transfer at their interfaces. Here, using a combined computational and experimental approach, we show that heterodyne vibrational sum frequency generation (HD-VSFG) measurements provide an effective way of monitoring the interfacial charge transfer in these heterostructures. Using ab initio quantum chemical calculations, we show that inducing regio-randomness into the organic polymer modifies the interfacial electronic states, the level of hybridization, and the electronic wave function of these materials. We present the HD-VSFG responses of the metal/P3HT heterojunctions containing both regio-regular and regio-random P3HT structures and show that the intensity of nonresonant signal is directly correlated with the computed electronic structure and the level of spontaneous charge transfer at these interfaces.