Complete NMR Assignment of MogrosidesII A2, II E andIII A1Isolated from Luo Han Guo

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Complete NMR Assignment of MogrosidesII… Table 2. 1 H and 13 C NMR (500 and 125 MHz, CD 3 OD), assignments forthe glycosides of 1, 2, and 3. 1 2 3 Position 13 C 1 H 13 C 1 H 13 C 1 H Glc I -1 --- --- 104.9 4.33 d (7.7) 104.2 4.43 d (7.3) Glc I -2 --- --- 75.3 3.21 m 81.2 3.61 m Glc I -3 --- --- 78.0 or 78.1 3.35 m 78.7 3.59 m or 78.3 Glc I -4 --- --- 71.6 or 71.7 3.28 m 71.6 3.33 m Glc I -5 --- --- 78.0 or 78.1 3.27 m 76.5 3.50 m or 78.3 Glc I -6 --- --- 62.6 3.64 m 3.84 m 70.1 3.62 m 4.23 dd (10.4, 1.8) Glc II -1 --- --- --- --- 104.4 or 104.5 4.28 d (7.7) Glc II -2 --- --- --- --- 75.2 3.20 m Glc II -3 --- --- --- --- 77.7-78.2 ¥ 3.36 m Glc II -4 --- --- --- --- 71.6 3.27 m Glc II -5 --- --- --- --- 77.7-78.2 ¥ 3.26 m Glc II -6 --- --- --- --- 62.7 3.65 m 3.85 m Glc III -1 --- --- --- --- 104.4 or 104.5 4.77 d (7.8) Glc III -2 --- --- --- --- 75.7 3.27 m Glc III -3 --- --- --- --- 77.7-78.2 ¥ 3.36 m Glc III -4 --- --- --- --- 72.4 3.21 m Glc III -5 --- --- --- --- 77.7-78.2 ¥ 3.27 m Glc III -6 --- --- --- --- 63.6 3.63 m 3.86 m Glc IV -1 106.4 4.28 d (7.8) 106.6 4.27 d (7.8) --- --- Glc IV -2 75.2 or 75.6 3.19 m 75.6 3.18 m --- --- Glc IV -3 78.1 3.31 m 78.0 or 78.1 3.31 m --- --- or 78.3 Glc IV -4 71.6 3.29 m 71.6 or 71.7 3.26 m --- --- Glc IV -5 77.2 3.40 m 77.7 3.21 m --- --- Glc IV -6 69.8 3.80 m 4.05 dd 62.8 3.65 m 3.81 m --- --- (11.8, 1.5) Glc V -1 104.8 4.42 d (7.8) --- --- --- --- Glc V -2 75.2 or 75.6 3.18 m --- --- --- --- Glc V -3 77.9 or 78.0 3.35 m --- --- --- --- Glc V -4 71.6 3.28 m --- --- --- --- Glc V -5 77.9 or 78.0 3.25 m --- --- --- --- Glc V -6 62.7 3.66 m --- --- --- --- 3.85 m ¥ Four carbon resonances in the range of 77.7-78.2 (77.66, 77.92, 77.05, and 78.16), hence chemical shifts could not be unequivocally assigned. The 1 H and 1 H- 13 C HSQC-DEPT data allowed identification of the two anomeric protons inMogroside II A 2 which were well resolved at H 4.42 ( C 104.8) and 4.28 ( C 106.4) and both protons had large couplings (7.8 Hz) indicating that they had β-configurations. The anomeric proton at H 4.28 showed an HMBC correlation to C-3 ( C 88.2)which indicated that it corresponded to Glc IV H-1.Glc IV H-2 and H-3 were then assigned based on COSY correlation between Glc IV H-1/Glc IV H-2 and Glc IV H-2/Glc IV H-3.Due to data overlap the COSY spectrum did not allow assignment of remaining Glc IV protons. Therefore, 1D TOCSY experiments were performed to assignGlc IV H-4, H-5 and H-6. HSQC-DEPT data was then used to assign the carbon chemical shift of Glc IV C-1 to C-6.HMBC correlations ofGlc IV H-1/C-2, C-3, C-5; H-2/C-1; H-3/C-2, C-4;H-4/C-6 and H- 6/C-4 further confirmed the assignments.The complete 1 H and 13 C assignments forGlc IV are presented in Table 2. The downfield carbon chemical shift of C-6 (δ C 69.8) indicated that the hydroxyl group at C-6 is replaced by a sugar linkage.The anomeric proton of Glc V at H 4.42 showed an HMBC correlation to the carbon at C 69.8 ppm (Glc IV C-6) and the reciprocal HMBC correlations from the methylene protons of Glc IV ( H 3.80 and 4.05) to the anomeric carbon ofGlc V at C 104.8 confirmedthe 16 linkage between Glc V and Glc IV .The remaining proton and carbon assignments for Glc V wasdone as discussed above for Glc IV . The key 1 H- 1 H COSY and 1 H- 13 C HMBCcorrelations for theglycone unit of Mogroside II A 2 (1) are presented in Figure 2 (B) and the complete proton and carbon assignments are presented in Table 2. 51
Complete NMR Assignment of MogrosidesII… Similarly in Mogroside II E(2),the presence of two anomeric protons were confirmed at H 4.33 ( C 104.9) and 4.27 ( C 106.6), having coupling of 7.7 Hz and 7.8 Hz, respectively, indicating that they hadconfigurations.The anomeric proton at H 4.27 showed an HMBC correlation to C-3( C 88.5)hence establishing the linkage ofGlc IV to C-3. The anomeric proton at H 4.33 showed an HMBC correlation to C-24 ( C 88.7)establishing the linkage ofGlc I to C-24. The COSY and 1D-TOCSY data allowed the assignments of Glc I and Glc IV H-2 to H-6 while the carbon assignments were based on HSQC-DEPT data and confirmed by HMBC correlations... The key 1 H- 1 H COSY and 1 H- 13 C HMBCcorrelations usedto assign the glucose units and their attachment to the triterpene core in Mogroside II E (2) are presented in Figure 3 and the complete proton and carbon assignments are presented in Table 2. Compound 2 is an isomer of compound 1, differing only in the position of the attachment of glucose units. In Mogroside III A 1 (3), the 1 H and HSQC-DEPT NMR data confirmed the presence of three glucose units.Theanomeric protons of these glucose units were observed at H 4.77 ( C 104.4 or 104.5), 4.43 ( C 104.2), and 4.28 ( C 104.4 or 104.5) and had large couplings (7.3 – 7.8 Hz) indicating that they had β-configurations. The anomeric proton at H 4.43 showed an HMBC correlation to C-24 ( C 93.3)which indicated that it corresponded to the anomeric proton of Glc I . The COSY and 1D-TOCSY experiments allowed the assignment of Glc I H-2 to Glc I H-6 and the HSQC-DEPT data was used to assign Glc I C-2 to Glc I C-6. HMBC correlations ofGlc I H-1/C-3; H-2/C-1, C-3; H-3/C-2; H-4/C-3, C-6 and H-5/C-6 further confirmed the assignments. The complete 1 H and 13 C assignments forGlc I are presented in Table 2. The downfield carbon chemical shifts of C-2 (δ C 81.2) and C-6 (δ C 70.1) indicated that the hydroxyl groups at C-2 and C-6 arereplaced by sugar linkages which was subsequently confirmed byHMBC correlations. The anomeric proton of Glc II at H 4.28 showed an HMBC correlation to the carbon at C 70.1 ppm (Glc I C-6) and the reciprocal HMBC correlation from the methylene protons of Glc I ( H 3.62 and 4.23) to the anomeric carbon ofGlc II at C 104.4 or 104.5 confirmed the 16 linkage between Glc II and Glc I . Similarly, HMBC correlation from the anomeric proton of Glc III at H 4.77 to the carbon at C 81.2 ppm (Glc I C-2) and the reciprocal HMBC correlation from the methine proton of Glc I H- 2 ( H 3.61) to the anomeric carbon ofGlc III at C 104.4 or 104.5 confirmed the 12 linkage between Glc III and Glc I .The assignment of remaining protons and carbons of Glc II and Glc III were done based on COSY, HSQC- DEPT, HMBC and 1D-TOCSY data. A summary of the key 1 H- 1 H COSY and 1 H- 13 C HMBCcorrelations of Mogroside III A 1 (3) used to assign the C-24 glycosides are described in Figure 4 and the complete 1 H and 13 C assignments are presented in Table 2. 52
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Complete NMR Assignment of MogrosidesII A2, II E andIII A1Isolated from Luo Han Guo

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