For a mechanical heart valve, a strong spike in pressure during closing is associated with valve wear and erythrocyte damage; thus, for valid in vitro testing, the mock circulation system should replicate the conditions, including pressure spikes, expected in vivo. To address this issue, a study was performed to investigate how mock circulation input impedance affects valve closure dynamics. A left ventricular model with polyurethane trileaflet inflow valve and tilting disc outflow valve was connected to a Louisville mock circulation system, which incorporates 2 adjustable flow resistors and 2 compliances. In the study, 116 cases matched zero frequency modulus well (982-1147 dyn x s/cm), but higher harmonics were purposely varied. Acceleration measured at the outflow valve ring (42.4-89.4 milli-Gs) was uncorrelated with impedance error (74.1-237 dyn x s/cm relative to target impedance), but was correlated with end-systolic impedance (1082-1319 dyn x s/cm) for cases with high zero frequency modulus, which exhibited just less than full ejection. These differences demonstrate that mock circulation response affects the magnitude of the closing spike, indicating that control of this parameter is necessary for authentic testing of valves. Correlation of acceleration to end-systolic impedance was weak for low zero frequency modulus, which tended toward full or hyperejection, reinforcing common laboratory observations that valve closing also depends on ventricular operating conditions.