## 3 Results and Discussion
### Hydrogenation of Limonin
From the \({}^{1}\)H-NMR data, it was observed that four methyl groups remained. Combined with the \({}^{13}\)C-NMR and DEPT spectral data, it was found that compounds **4** and **5** had one ester carbonyl group, five methylene groups, and seven quaternary carbons. Compared with compound **2**, one ester carbonyl group was lost in **4** and **5**. It could be inferred that the ester carbonyl group on ring A or ring B was reduced to a hydroxyl group. According to the 2D-NMR spectra of \({}^{1}\)H-\({}^{1}\)H Correlated Spectroscopy (COSY) (Figures S16 and S17), HSQC (Figures S18 and S19), and HMBC (Figures S20 and S21), the carbonyl groups at the C-3 or C-16 position were also reduced. The configuration of these hydroxyl groups was then determined by the NOESY spectrum (Figures S22 and S23). First, C-20 (\(\delta_{\rm C}\) 124.83) correlated with H-17 (\(\delta_{\rm H}\) 4.97), confirming the hydrogen shifted on C-17 (\(\delta_{\rm C}\) 73.77). It was observed by the HMBC spectrum that H-17 was correlated with \(\delta_{\rm C}\) 38.38, \(\delta_{\rm C}\) 21.91, \(\delta_{\rm C}\) 68.65, and \(\delta_{\rm C}\) 89.07, respectively. In addition, \(\delta_{\rm C}\) 68.65 and \(\delta_{\rm C}\) 89.07 were attached to oxygen atoms, in which \(\delta_{\rm C}\) 68.65 was quaternary carbon, and \(\delta_{\rm C}\) 89.07 was methane. Thus, \(\delta_{\rm C}\) 68.65 was C-14, and \(\delta_{\rm C}\) 89.07 was C-16. According to \({}^{1}\)H-1H COSY, H-15 (\(\delta_{\rm H}\) 3.96) correlated with H-16 (\(\delta_{\rm H}\) 5.30). Moreover, H-1 (\(\delta_{\rm H}\) 4.06) and H-19 (\(\delta_{\rm H}\) 4.46, 4.63) were correlated with C-3 (\(\delta_{\rm C}\) 173.73), respectively. The relative configurations of compounds **4** and **5** were different. By analyzing their NOESY spectra (Figures S24 and S25) and the three-dimensional model, the relative configuration issues were primarily caused by the hydroxyl groups at C-7 and C-16 positions. Due to its rather rigid structure, limonin did not change the configuration at positions other than the reaction sites, such as H-1, H-4, H-9, and C-24. In the NOESY spectrum, H-5 (  
Figure 4: Hydrogenation of limonin by lithium aluminum hydride. 1.99), H-7 (\(\delta_{\rm H}\) 3.94), and H-9 (\(\delta_{\rm H}\) 2.36) were correlated with each other, respectively; the methyl groups of H-16, H-15, and C-18 (\(\delta_{\rm H}\) 1.29) also showed NOE correlation with each other, respectively. Based on the above two groups of NOE correlations, both C-7 and C-16 positions of compound **4** were \(\beta\)-OH. The NOESY spectrum of compound **5** clearly showed the NOE correlation between H-7 and H-24 (\(\delta_{\rm H}\) 0.80), as well as the correlation between H-5 and H-9. In contrast, no NOE correlation with H-7 was seen. This proved that the C-7 position was \(\alpha\)-OH, and also explained the difference in chemical shift around it. Furthermore, H-16 (\(\delta_{\rm H}\) 5.27), H-15 (\(\delta_{\rm H}\) 3.48), and H-18 (\(\delta_{\rm H}\) 1.28) also had NOE correlations with each other, respectively, confirming that compounds **5** and **4** were \(\beta\)-OH at C-16 position. Compounds **6** and **7** were formed under the same conditions. They were obtained by the subsequent hydrogenation using sodium borohydride for 90 min on the basis of compounds **4** and **5**. The HR-ESI-MS results indicated that the two compounds shared the same \(m\)/\(z\) signal of 479.2644. Combined with the \({}^{1}\)H and \({}^{13}\)C NMR data (Figures S26-S29), it was deduced that they were isomers with the molecular formula of C\({}_{\rm 26}\)H\({}_{\rm 38}\)O\({}_{\rm s}\), and the degree of unsaturation of eight. According to \({}^{13}\)C-NMR, DEPT, and HSQC spectra (Figures S30 and S31) analysis, compound **6** had six quaternary carbons, six methylene groups, 10 methine groups, and four methyl groups. Combined with the infrared spectrum analysis, no signal of ester carbonyl was observed. It was deduced that one of its rings was opened.