Ultrasonic has been used since several decades ago. One of ultrasonic application is used in medical field to observe mechanical properties of target, such as speed of sound, density, acoustic impedance, attenuation, etc. Several methods are also available to estimate mechanical property of target. In the previous research, we developed Scanning Acoustic Microscope (SAM). On that research, we proposed a method to estimated acoustic impedance by utilizing calibration curve. A calibration curve is established based on echo intensity obtained from calculation of acoustic propagation. The highest amplitude of a single frequency component is chosen and taken into calculation. The other frequency components are neglected. In this paper, authors propose a method to estimate acoustic impedance of target. The acoustic impedance is estimated based on calibration curve by considering all frequency components of acoustic wave. This calculation is then compared to previous research. As a result, acoustic impedance of cerebellar tissue of rat are presented by utilizing two calibration curves.