Objective To study changes and correlations in ultrasound acoustic parameters, bone density and microstructure of the cancellous bone at different stages of decalcification. Methods Fifteen defatted porcine cancellous bone specimens were decalcified at different decalcification stages, and the bone density, microstructure and acoustic parameters were measured by Micro CT and ultrasound system, respectively, before and after the decalcification. Correlations between acoustic parameters, bone density and microstructure were investigated. Results With the loss of calcium in bone specimens, BMD (bone mass density), BS/TV and BV/TV decreased continuously. Microstructure parameters SMI and BS/BV increased, while Tb.Th and Tb.N decreased with Tb. Sp increasing. Degree of anisotropy (DA) increased. Acoustic parameter SOS increased at first, and then decreased, with nBUA slightly decreasing. High correlation was found between acoustic parameters, BMD and bone microstructure parameters. Conclusions Ultrasound acoustic parameters are correlated with BMD and bone microstructure. This study may provide some reference information for the early diagnosis of osteoporosis based on ultrasound.