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Purpose  Microscopic fractional anisotropy (µFA) can disentangle microstructural information from orientation dispersion. While double diffusion encoding (DDE) MRI methods are widely used to extract accurate µFA, it has only recently been proposed that powder‐averaged single diffusion encoding (SDE) signals, when coupled with the diffusion standard model (SM) and a set of constraints, could be used for µFA estimation. This study aims to evaluate µFA as derived from the spherical mean technique (SMT) set of constraints, as well as more generally for powder‐averaged SM signals.    Methods  SDE experiments were performed at 16.4 T on an ex vivo mouse brain (Δ/δ = 12/1.5 ms). The µFA maps obtained from powder‐averaged SDE signals were then compared to maps obtained from DDE‐MRI experiments (Δ/τ/δ = 12/12/1.5 ms), which allow a model‐free estimation of µFA. Theory and simulations that consider different types of heterogeneity are presented for corroborating the experimental findings.    Results  µFA, as well as other estimates derived from powder‐averaged SDE signals produced large deviations from the ground truth in both gray and white matter. Simulations revealed that these misestimations are likely a consequence of factors not considered by the underlying microstructural models (such as intercomponent and intracompartmental kurtosis).    Conclusion  Powder‐averaged SMT and (2‐component) SM are unable to accurately report µFA and other microstructural parameters in ex vivo tissues. Improper model assumptions and constraints can significantly compromise parameter specificity. Further developments and validations are required prior to implementation of these models in clinical or preclinical research.

Microscopic anisotropy misestimation in spherical‐mean single diffusion encoding MRI