Ultrasonic gas body activation in Elodea leaves and the mechanical index

Membrane damage resulting from ultrasonic gas body activation was investigated in leaves of the aquatic plant Elodea using pulse-mode exposures from 0.745-15 MHz. The frequency response was similar to that previously observed for continuous exposures. Cell death thresholds were higher for the pulse modes; for example, at 6 MHz the threshold was 166 W/cm2 spatial-peak, pulse-average (SPPA) intensity for 1 microsecond pulses and 1 kHz pulse repetition frequency (PRF) for 60 s exposures, compared to 12 W/cm2 for 60 ms continuous exposure. Increasing the PRF for 2.5 MHz, 3 microseconds pulses from 10 Hz to 5 kHz resulted in a gradual decrease in the threshold. Increasing the pulse duration from 1 microsecond to 30 microseconds and PRF from 33 Hz to 1 kHz only weakly influenced the threshold, for the constant 60 ms total on-time. Decreasing the pulse duration for single 6 MHz pulses increased the threshold up to 1,540 W/cm2 at 30 microseconds. The gas body activity was modeled with linear theory for oscillation of the gas channel walls, and intracellular shear stress, which causes lethal damage to the plasma membranes, was modeled with theory for acoustic microstreaming. Theory and observation indicate that the minimum shear stress threshold as a function of resonance frequency has essentially the same form as the Mechanical Index (MI) in the 0.5 to 20 MHz range. Specifically, for 3 microseconds pulses with 0.2-2 kHz PRF and 60 ms total on-time, the pressure-amplitude threshold divided by the square root of the frequency was approximately constant at 0.95 MPa/MHz1/2.