Isoxazolidines serve as intermediates in the synthesis of natural products. In addition, they can display significant biological activity, much of which derives from their ability to act as nucleoside analogues. As a result, considerable effort has been applied toward the asymmetric synthesis of isoxazolidines. However, a rapid and straightforward method for determination of the absolute configuration of isoxazolidines has not yet been reported. Herein we report the application of Mosher derivatives for the determination of the absolute configuration of substituted isoxazolidines. The Mosher derivatives exhibit conformational behavior similar to the Mosher derivatives of cyclic secondary amines. Interpretation of the chemical shift anisotropy, with these conformational biases in mind, can be used for the assignment of the absolute configuration of substituted isoxazolidines.