The ultrasonic absorption coefficient per wavelength (αλ), as a function of temperature and frequency, was determined for large unilamellar vesicles (LUV) in the vicinity of their phospholipid phase transition temperature, using a double crystal acoustic interferometer. (The vesicles were composed of a 4:1 (w/w) mixture of dipalmitoylphosphatidylcholine (DPPC) and dipalmitoylphosphatidylglycerol (DPPG)). It has been found that αλ reaches a maximum ( αλ) max at the phase transition temperature ( t m) of the phospholipids in the bilayer, at an ultrasonic relaxation frequency of 2.1 MHz. Divalent cations (Ca 2+ and Mg 2+), added to LUV suspensions, shifted ( αλ) max to higher temperatures, dependent upon the concentration of divalent cation. It was also found that the shape of the αλ versus t curve was significantly changed, representing changes in the Van't Hoff enthalpy of the transition, and therefore, the cooperative unit of the transition. This suggests that divalent cations interact individually with the negatively charged phospholipid headgroups of DPPG and with DPPC headgroups, thus decreasing the cooperative unit of the transition. The observed upward shift in t m suggests an interaction that increases the activation energy and, therefore, the temperature of the phase transition. However, αλ as a function of frequency did not change with the addition of divalent cations and, thus, the relaxation time of the event responsible for the absorption of ultrasound is not changed by the addition of divalent cations.