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This paper presents a new method to experimentally characterize the rotational self-loosening behavior of bolted joints. Furthermore, a method to calculate the preload loss due to certain relative displacements of the clamped parts was developed. The method obtains all required calculation parameters from the experimental tests performed and neither needs further experimental determination of friction coefficients nor numerical simulations to predict the preload loss. The method accumulates the preload loss of load cycles. Thereby, it can be used to estimate the time a machine can operate until a critical amount of preload is lost. Non-linear dependencies of the preload loss to the displacement amplitude and the remaining preload are considered. The calculation method also applies to load cases with variable load amplitudes. This is validated experimentally.