A novel pyrrolizidine alkaloid was isolated and purified from the aerial parts of Crotalaria re-tusa L. Structural elucidation of the alkaloid was established by analyses of high field NMR spectra and quantum mechanical calculations. The H-1 and C-13 NMR chemical shifts for eight candidate structures were calculated and compared with the experimental NMR data applying the corrected mean absolute error (CMAE) and the statistical probability methodology (DP4). The interpretation of the experimental NMR spectra and CMAE and DP4 analyses established that the novel pyrrolizidine alkaloid is (3S*, 4R*, 5S*, 8a(1)R*, 13aR*)-3-hydroxy-3,4,5-trimethyl-4,5,8,8a(1),10,12,13,13-aoctahydro-2H- [1,6]dioxacycloundecino [2,3,4 gh]pyrrolizine-2,6(3H)-dione. (C) 2022 Elsevier B.V. All rights reserved.