Table 2.
Structural identification of four oxygenated 13R-manoyl oxide-derived diterpenoids formed following transient expression of CYP encoding genes from C. forskohlii based on comparison of their 1H-NMR and 13C-NMR (Figure 4—source data 1) chemical shifts to literature data. Chemical shifts for reference compounds marked with * have not been assigned to a specific carbon. The 13C chemical shifts of 9-deoxyforskolin (Gabetta et al., 1989) were used as reference for 6,7-dihydroxy-11-oxo-13R-manoyl oxide (7h).
| 9-Deoxydeacetylforskolin (10b)† | 1,9-Dideoxydeacetylforskolin (7h)† | 11-oxo-13R-manoyl oxide (2)† | Coleorol (3a)† | |||||||
|---|---|---|---|---|---|---|---|---|---|---|
| Pos. | 1H (nH; m; J(Hz)) | 13C | (Gabetta et al., 1989) | 1H (nH; m; J(Hz)) | 13C | (Gabetta et al., 1989) | 13C | (Gabetta et al., 1989) | 13C | (Asada et al., 2012) |
| 1 | 4.38 (1H; t; 2.8) | 71.6 | 71.2 | 2.45 (1H, d(br); 13.1) 0.78 (H; m) |
41.5 | 43.1 | 42.1 | 41.9 | 31.7 | 31.6 |
| 2 | 1.47 (1H; m) 2.14 (1H; m) |
25.8 | 25.6 | 1.78 (H; m) 1.40 (H; m) |
18.7 | 18.4 | 18.5 | 18.4 | 18.6 | 18.4 |
| 3 | 1.12 (1H; dt; 3.4, 13.2) 1.62 (1H; dt; 3.5, 13.5) |
36.4 | 36.3 | 1.36 (H; m) 1.15 (H; m) |
43.8 | 43.7 | 43.4 | 43.3 | 41.9 | 41.8 |
| 4 | 34.2 | 34.1 | 34.4 | 34.1 | 33.4 | 33.2 | 33.3 | 33.2 | ||
| 5 | 1.34 (1H; d; 2.1) | 47.5 | 47.4 | n.d. | 55.7 | 55.2 | 56.0 | 55.8 | 45.7 | 45.5 |
| 6 | 4.44 (1H; t; 2.6) | 70.8 | 70.2 | 4.39 (1H; m) | 70.4 | 70.2 | 19.8 | 19.7 | 19.5 | 19.4 |
| 7 | 3.68 (1H; d; 3.6) | 80.7 | 81.1 | 3.71 (1H; d; 3.8) | 81.0 | 80.7 | 39.6 | 39.4 | 36.6 | 36.4 |
| 8 | 80.0 | 78.5 | 80.1 | 79.9 | 77.5 | 77.2 | 78.0 | 77.8 | ||
| 9 | 3.32 (1H; s) | 58.0 | 58.2 | 2.59 (1H; s) | 65.5 | 65.4 | 66.9 | 66.7 | 75.3 | 75.2 |
| 10 | 42.2 | 41.7 | 38.0 | 37.8 | 37.3 | 37.1 | 41.1 | 40.9 | ||
| 11 | 207.7 | 207.6 | 206.3 | 205.7 | 207.7 | 207.1 | 21.1 | 21.0 | ||
| 12 | 2.63 (1H; d; 18.0) 2.69 (1H; d; 18.0) |
49.8 | 49.9 | 2.60 (1H; d; 18.1) 2.66 (1H; d; 18.1) |
50.0 | 49.8 | 50.4 | 50.2 | 31.6 | 31.5 |
| 13 | 75.1 | 74.8 | 75.1 | 75.1 | 75.1 | 74.4 | 72.9 | 72.8 | ||
| 14 | 5.94 (1H; dd; 10.8, 17.4) | 146.2 | 145.8 | 5.95 (1H; dd; 10.7, 17.4) | 146.9 | 146.4 | 146.9 | 146.0 | 147.4 | 147.3 |
| 15 | 5.04 (1H; d; 10.8) 5.14 (1H; d; 17.4) |
112.4 | 112.7 | 5.04 (1H; d; 10.7) 5.17 (1H; d; 17.4) |
112.3 | 112.1 | 112.3 | 111.9 | 110.1 | 110.0 |
| 16 | 1.30 (3H; s) | 31.5 | 31.5* | 1.28 (3H; s) | 31.6 | 33.2* | 31.4 | 31.2* | 28.9 | 28.8 |
| 17 | 1.54 (3H; s) | 24.1 | 24.5* | 1.50 (3H; s) | 23.5 | 31.4* | 28.1 | 27.9* | 27.0 | 29.9 |
| 18 | 1.38 (3H; s) | 33.1 | 18.2* | 0.97 (3H; s) | 33.4 | 23.9* | 15.6 | 15.5* | 33.7 | 33.6 |
| 19 | 1.21 (3H; s) | 23.7 | 23.6* | 1.21 (3H; s) | 24.0 | 23.7* | 21.8 | 21.6* | 21.5 | 21.4 |
| 20 | 1.01 (3H; s) | 18.5 | 32.8* | 1.30 (3H; s) | 17.2 | 16.7* | 33.6 | 33.5* | 17.0 | 16.8 |
†1H and 13C NMR data acquired at 600 and 150 MHz, respectively,in methanol-d4, at 300 K. s = singlet, d = doublet, t = triplet, m = multiplet, br = broad.