Amines are one of the common functional groups of interest due to their abundant presence in natural proteins, surfactants and other chemicals. However, their accurate spectral assignment of vibrational modes, critical to interpreting SFG signals for characterizing various bio-interfaces such as protein–membrane interaction and surfactant adsorption, still remains elusive. Herein we present a systematic study to identify and justify the correct peak assignment of the N⁺–H stretching mode at the air–water interface. We used three special surfactants: hexadecylamine (a primary amine without counterions), dodecylamine hydrochloride (a primary amine with counterions) and hexadecyltrimethylammonium bromide as a control (the N⁺–H stretching mode is absent in this quarternary amine). We suppressed the SFG interfacial water signals using saturated NaCl solutions. Our designed experiments resolved the current controversy and concluded that the 3080 cm⁻¹ peak is from the N⁺–H vibrations, while the 3330 cm⁻¹ peak is not due to ammonium species but rather originates from the interfacial water vibrational modes or the backbone amide modes.