In designing acoustic detection systems for stored grain state, acoustic attenuation coefficient of stored grain, that is, the amount of attenuation of sound waves due to scattering and absorption in the propagation path per unit length is a crucial knowledge. When the sound source emits a single tone signal, it comes down to the problem of the amplitude estimation of a sinusoidal signal. A measurement method of acoustic attenuation coefficient based on orthogonal sequence demodulation is proposed. A measurement system of acoustic attenuation coefficient is developed. The sound pressure amplitude at the near and far microphones was estimated by orthogonal sequence demodulation, and then the acoustic attenuation coefficient can be calculated. Test results show that the attenuation ratio acquired by the proposed method is accurate and stable. Using the system, the curve of the attenuation coefficient of the sound wave in soybean varying with frequency can be obtained easily.