3.3. Acetylation
Fatsicarpain D (2.0 mg) was dissolved in pyridine (0.5 mL) and allowed to react overnight at room temperature with acetic anhydride (one drop). Then, the reaction was quenched by the addition of 1.0 mL of H2O, followed by extraction with EtOAc (3 × 1.0 mL). The EtOAc extracts were combined and evaporated. The resulting residue was subjected to a short silica gel column using n-hexane–EtOAc (3:1) to yield an acetylated product 1 (1.9 mg). Additionally, fatsicarpain C (3.0 mg), 3α-hydroxyolean-11-en-28,13β-olide (3.0 mg), fatsicarpain F (3.0 mg) and 3α-hydroxyolean-11,13(18)-dien-28-oic acid (3.0 mg) were subjected to acetylation, and chromatographic purification of the product according to the same procedure mentioned under acetylation of fatsicarpain D, to give 2 (2.9 mg), 3 (2.8 mg), 4 (2.7 mg) and 5 (2.9 mg), respectively.
3α-Acetoxyolean-9,12-dien-28-oic acid (1). Yield 91%. white amorphous powder; [α]25D +75 (c = 0.1, CHCl3); UV (MeOH) λmax (log ε) 212 (4.40), 274 (3.85) nm; IR (KBr) νmax 3,458, 2,941, 1,733, 1,700, 1,455, 1,374, 1,245, 758 cm−1; 1H-NMR (CDCl3, 600 MHz) δH 1.59 (1H, m, H-1α), 1.76 (1H, m, H-1β), 1.73 (1H, m, H-2α), 1.94 (1H, m, H-2β), 4.64 (1H, br t, J = 3.0 Hz, H-3), 1.31 (1H, m, H-5), 1.50 (2H, m, H-6), 1.72 (1H, m, H-7α), 1.32 (1H, m, H-7β), 5.60 (1H, d, J = 6.0 Hz, H-11), 5.58 (1H, d, J = 6.0 Hz, H-12), 1.16 (1H, m, H-15α), 1.83 (1H, m, H-15β), 1.99 (1H, m, H-16α), 1.68 (1H, m, H-16β), 2.98 (1H, dd, J = 13.8, 4.2 Hz, H-18), 1.62 (1H, m, H-19α), 1.20 (1H, m, H-19β), 1.23 (1H, m, H-21α), 1.36 (1H, m, H-21β), 1.79 (1H, m, H-22α), 1.66 (1H, m, H-22β), 0.89 (3H, s, H-23), 0.91 (1H, s, H-24), 1.19 (3H, s, H-25), 0.99 (3H, s, H-26), 1.08 (3H, s, H-27), 0.95 (3H, s, H-29), 0.90 (3H, s, H-30), 2.04 (3H, s, 3-OAc); 13C-NMR (CDCl3, 150 MHz) δC 32.2 (t, C-1), 23.3 (t, C-2), 77.8 (d, C-3), 36.8 (s, C-4), 46.0 (d, C-5), 18.0 (t, C-6), 31.8 (t, C-7), 42.4 (s, C-8), 154.8 (s, C-9), 38.7 (s, C-10), 115.2 (d, C-11), 120.7 (d, C-12), 144.7 (s, C-13), 40.8 (s, C-14), 26.9 (t, C-15), 23.6 (t, C-16), 45.9 (s, C-17), 39.4 (d, C-18), 45.9 (t, C-19), 30.6 (s, C-20), 33.7 (t, C-21), 32.1 (t, C-22), 27.8 (q, C-23), 21.9 (q, C-24), 24.8 (q, C-25), 20.4 (q, C-26), 20.1 (q, C-27), 182.8 (s, C-28), 23.5 (q, C-29), 32.9 (q, C-30), 170.8 (s, 3-OAc), 21.3 (q, 3-OAc); ESIMS m/z 519 [M+Na]+; HRESIMS m/z 519.3445 [M+Na]+ (Calcd for C32H48O4Na, 519.3445).
3α-Acetoxy-24-formylolean-11,13(18)-dien-28-oic acid (2). Yield 85%. white amorphous powder; [α]25D −64 (c = 0.1, CHCl3); UV (MeOH) λmax (log ε) 235 (4.08), 247 (3.78), 263 (3.55) nm; IR (KBr) νmax 3,454, 2,941, 2,861, 1,734, 1,698, 1,450, 1,373, 1,242, 759 cm−1; 1H-NMR (CDCl3, 600 MHz) δH 1.33 (1H, m, H-1α), 1.51 (1H, m, H-1β), 1.78 (1H, m, H-2α), 1.93 (1H, m, H-2β), 4.87 (1H, br s, H-3), 2.01 (1H, m, H-5), 1.50 (2H, m, H-6), 1.33 (1H, m, H-7α), 1.25 (1H, m, H-7β), 2.20 (1H, br s, H-9), 5.64 (1H, d, J = 10.2 Hz, H-11), 6.47 (1H, dd, J = 10.2, 3.0 Hz, H-12), 1.10 (1H, m, H-15α), 1.70 (1H, m, H-15β), 2.01 (1H, m, H-16α), 1.71 (1H, m, H-16β), 2.54 (1H, d, J = 14.4 Hz, H-19α), 1.69 (1H, d, J = 14.4 Hz, H-19β), 1.32 (1H, m, H-21α), 1.39 (1H, m, H-21β), 2.28 (1H, m, H-22α), 1.40 (1H, m, H-22β), 1.08 (3H, s, H-23), 9.41 (1H, s, H-24), 0.97 (3H, s, H-25), 0.82 (3H, s, H-26), 1.06 (3H, s, H-27), 0.81 (3H, s, H-29), 0.96 (3H, s, H-30), 2.06 (3H, s, 3-OAc); 13C-NMR (CDCl3, 150 MHz) δC 31.8 (t, C-1), 22.2 (t, C-2), 74.6 (d, C-3), 51.1 (s, C-4), 43.9 (d, C-5), 20.3 (t, C-6), 31.9 (t, C-7), 41.2 (s, C-8), 53.9 (d, C-9), 36.1 (s, C-10), 126.2 (d, C-11), 125.5 (d, C-12), 136.8 (s, C-13), 42.2 (s, C-14), 24.8 (t, C-15), 32.6 (t, C-16), 48.0 (s, C-17), 131.5 (s, C-18), 40.5 (t, C-19), 32.4 (s, C-20), 36.8 (t, C-21), 35.4 (t, C-22), 14.0 (q, C-23), 205.9 (d, C-24), 17.8 (q, C-25), 16.6 (q, C-26), 19.9 (q, C-27), 180.9 (s, C-28), 24.0 (q, C-29), 32.2 (q, C-30), 170.0 (s, 3-OAc), 21.2 (q, 3-OAc); ESIMS m/z 533 [M + Na]+; HRESIMS m/z 533.3207 [M + Na]+ (Calcd for C32H46O5Na, 533.3237).
3α-Acetoxyolean-11-en-28,13β-olide (3). Yield 82%. white amorphous powder; [α]25D +52 (c = 0.1, CHCl3); IR (KBr) vmax 2,941, 1,750, 1,736, 1,456, 1,376, 1,247, 1,158, 756 cm−1; 1H-NMR (CDCl3, 600 MHz) δH 1.17 (1H, m, H-1α), 1.62 (1H, m, H-1β), 1.63 (1H, m, H-2α), 1.95 (1H, m, H-2β), 4.65 (1H, t, J = 2.4 Hz, H-3), 1.24 (1H, m, H-5), 1.50 (2H, m, H-6), 1.45 (1H, m, H-7α), 1.26 (1H, m, H-7β), 2.01 (1H, br s, H-9), 6.05 (1H, d, J = 10.2 Hz, H-11), 5.42 (1H, dd, J = 10.2, 3.0 Hz, H-12), 1.21 (1H, m, H-15α), 1.73 (1H, m, H-15β), 2.13 (1H, td, J = 13.2, 6.6 Hz, H-16α), 1.36 (1H, m, H-16β), 2.08 (1H, m, H-18), 1.82 (1H, t, J = 13.2 Hz, H-19α), 1.36 (1H, m, H-19β), 1.32 (1H, m, H-21α), 1.36 (1H, m, H-21β), 1.65 (1H, m, H-22α), 1.69 (1H, m, H-22β), 0.85 (3H, s, H-23), 0.90 (3H, s, H-24), 0.93 (3H, s, H-25), 1.07 (3H, s, H-26), 1.12 (3H, s, H-27), 0.88 (3H, s, H-29), 0.97 (3H, s, H-30), 2.08 (3H, s, 3-OAc); 13C-NMR (CDCl3, 150 MHz) δC 33.5 (t, C-1), 22.6 (t, C-2), 78.1 (d, C-3), 36.7 (s, C-4), 49.6 (d, C-5), 17.4 (t, C-6), 30.9 (t, C-7), 41.4 (s, C-8), 52.9 (d, C-9), 36.4 (s, C-10), 135.8 (d, C-11), 126.8 (d, C-12), 89.8 (s, C-13), 41.7 (s, C-14), 25.3 (t, C-15), 21.3 (t, C-16), 44.0 (s, C-17), 50.5 (d, C-18), 37.3 (t, C-19), 31.4 (s, C-20), 34.3 (t, C-21), 27.1 (t, C-22), 27.6 (q, C-23), 21.2 (q, C-24), 17.7 (q, C-25), 19.0 (q, C-26), 18.5 (q, C-27), 180.1 (s, C-28), 23.5 (q, C-29), 33.2 (q, C-30), 170.7 (s, 3-OAc), 21.4 (q, 3-OAc); ESIMS m/z 519 [M + Na]+; HRESIMS m/z 519.3422 [M + Na]+ (Calcd for C32H48O4Na, 519.3445).
3α-Acetoxy-11α-methoxyolean-12-en-28-oic acid (4). Yield 83%. white amorphous powder; [α]25D −76 (c = 0.1, CHCl3); IR (KBr) vmax 2,941, 1,737, 1,697, 1,450, 1,373, 1,243, 1,110, 756 cm−1; 1H-NMR (CDCl3, 600 MHz) δH 1.49 (1H, m, H-1α), 1.60 (1H, m, H-1β), 1.61 (1H, m, H-2α), 2.06 (1H, m, H-2β), 4.62 (1H, br s, H-3), 1.24 (1H, m, H-5), 1.46 (2H, m, H-6), 1.50 (1H, m, H-7α), 1.22 (1H, m, H-7β), 1.79 (1H, d, J = 8.4 Hz, H-9), 3.74 (1H, dd, J = 8.4, 3.6 Hz, H-11), 5.53 (1H, d, J = 3.6 Hz, H-12), 1.18 (1H, m, H-15α), 2.01 (1H, m, H-15β), 2.03 (1H, m, H-16α), 1.63 (1H, m, H-16β), 2.87 (1H, dd, J = 13.2, 3.6 Hz, H-18), 1.61 (1H, m, H-19α), 1.19 (1H, m, H-19β), 1.24 (1H, m, H-21α), 1.37 (1H, m, H-21β), 1.80 (1H, m, H-22α), 1.62 (1H, m, H-22β), 0.85 (3H, s, H-23), 0.89 (3H, s, H-24), 1.03 (3H, s, H-25), 0.78 (3H, s, H-26), 1.26 (3H, s, H-27), 0.96 (3H, s, H-29), 0.93 (3H, s, H-30), 2.07 (3H, s, 3-OAc); 13C-NMR (CDCl3, 150 MHz) δC 34.3 (t, C-1), 29.4 (t, C-2), 78.4 (d, C-3), 36.5 (s, C-4), 49.9 (d, C-5), 18.1 (t, C-6), 32.8 (t, C-7), 42.8 (s, C-8), 53.0 (d, C-9), 38.2 (s, C-10), 76.0 (d, C-11), 121.6 (d, C-12), 148.3 (s, C-13), 41.9 (s, C-14), 27.8 (t, C-15), 22.8 (t, C-16), 46.0 (s, C-17), 40.7 (d, C-18), 45.6 (t, C-19), 30.7 (s, C-20), 33.7 (t, C-21), 32.2 (t, C-22), 28.0 (q, C-23), 21.9 (q, C-24), 16.8 (q, C-25), 18.7 (q, C-26), 25.5 (q, C-27), 180.4 (s, C-28), 23.5 (q, C-29), 33.0 (q, C-30), 170.9 (s, 3-OAc), 21.4 (q, 3-OAc); ESIMS m/z 551 [M + Na]+; HRESIMS m/z 551.3686 [M + Na]+ (Calcd for C33H52O5Na, 551.3707).
3α-Acetoxy olean-11,13(18)-dien-28-oic acid (5). Yield 85%. white amorphous powder; [α]25D −51 (c = 0.1, CHCl3); UV (MeOH) λmax (log ε) 232 (4.10), 245 (3.68), 264 (3.65) nm; IR (KBr) vmax 3,464, 2,941, 1,731, 1,696, 1,456, 1,376, 1,247, 756 cm−1; 1H-NMR (CDCl3, 600 MHz) δH 1.25 (1H, m, H-1α), 1.64 (1H, m, H-1β), 1.64 (1H, m, H-2α), 1.96 (1H, m, H-2β), 4.66 (1H, br s, H-3), 1.30 (1H, m, H-5), 1.50 (2H, m, H-6), 1.38 (1H, m, H-7α), 1.34 (1H, m, H-7β), 2.05 (1H, br s, H-9), 5.65 (1H, d, J = 10.2 Hz, H-11), 6.43 (1H, dd, J = 10.2, 2.4 Hz, H-12), 1.09 (1H, m, H-15α), 1.72 (1H, m, H-15β), 2.00 (1H, m, H-16α), 1.72 (1H, m, H-16β), 2.54 (1H, d, J = 14.4 Hz, H-19α), 1.67 (1H, d, J = 14.4 Hz, H-19β), 1.29 (1H, m, H-21α), 1.40 (1H, m, H-21β), 2.27 (1H, m, H-22α), 1.40 (1H, m, H-22β), 0.89 (3H, s, H-23), 0.85 (3H, s, H-24), 0.93 (3H, s, H-25), 0.80 (3H, s, H-26), 1.02 (3H, s, H-27), 0.81 (3H, s, H-29), 0.95 (3H, s, H-30), 2.09 (3H, s, 3-OAc); 13C-NMR (CDCl3, 150 MHz) δC 33.3 (t, C-1), 22.6 (t, C-2), 78.3 (d, C-3), 36.7 (s, C-4), 49.7 (d, C-5), 18.0 (t, C-6), 32.2 (t, C-7), 40.8 (s, C-8), 54.1 (d, C-9), 36.7 (s, C-10), 127.0 (d, C-11), 125.2 (d, C-12), 137.0 (s, C-13), 42.0 (s, C-14), 24.8 (t, C-15), 32.6 (t, C-16), 48.0 (s, C-17), 130.9 (s, C-18), 40.4 (t, C-19), 32.5 (s, C-20), 36.8 (t, C-21), 35.5 (t, C-22), 27.7 (q, C-23), 21.3 (q, C-24), 17.8 (q, C-25), 16.4 (q, C-26), 19.9 (q, C-27), 181.9 (s, C-28), 24.0 (q, C-29), 32.2 (q, C-30), 170.9 (s, 3-OAc), 21.4 (q, 3-OAc); ESIMS m/z 519 [M + Na]+; HRESIMS m/z 519.3422 [M+Na]+ (Calcd for C32H48O4Na, 519.3445).
3.4. Acylation
To a solution of fatsicarpain A (5.0 mg) in CH2Cl2 (1.0 mL) EDC·HCl (1.0 mg) was added. The solution was stirred for 3 h at 50 °C. After the completion of the reaction the solvent was evaporated under reduced pressure to give a crude product which was subjected to a short silica gel column eluting with n-hexane–EtOAc (1:1) to yield 6 (3.2 mg).
N-(3-(Dimethylamino)propyl)-N-(ethylcarbamoyl)-3α,23-dihydroxyolean-11,13(18)-dien-28-amide (6). Yield 48%. white amorphous powder; [α]25D –13 (c = 0.1, CHCl3); UV (MeOH) λmax (log ε) 236 (4.11), 247 (3.88), 262 (3.74) nm; IR (KBr) vmax 3,414, 2,933, 1,696, 1,686, 1,651, 1,463, 1,363, 1,256, 1,225, 1,051, 756 cm−1; 1H-NMR (CDCl3, 600 MHz) δH 1.39 (1H, m, H-1α), 1.64 (1H, m, H-1β), 1.53 (1H, m, H-2α), 2.02 (1H, m, H-2β), 3.72 (1H, br s, H-3), 1.74 (1H, m, H-5), 1.45 (2H, m, H-6), 1.42 (2H, m, H-7), 2.14 (1H, br s, H-9), 5.65 (1H, d, J = 10.2 Hz, H-11), 6.43 (1H, d, J = 10.2, 3.0 Hz, H-12), 1.07 (1H, m, H-15α), 1.65 (1H, m, H-15β), 2.63 (1H, m, H-16α), 1.64 (1H, m, H-16β), 2.53 (1H, d, J = 14.4 Hz, H-19α), 1.47 (1H, d, J = 14.4 Hz, H-19β), 1.28 (1H, m, H-21α), 1.42 (1H, m, H-21β), 2.41 (1H, m, H-22α), 1.26 (1H, m, H-22β), 3.55 (1H, d, J = 11.4 Hz, H-23a), 3.42 (1H, d, J = 11.4 Hz, H-23b), 0.70 (3H, s, H-24), 0.95 (3H, s, H-25), 0.89 (3H, s, H-26), 1.00 (3H, s, H-27), 0.83 (3H, s, H-29), 0.92 (3H, s, H-30), 4.08 (1H, m, H-1'a), 3.38 (1H, m, H-1'b), 2.02 (1H, m, H-2'a), 1.75 (1H, m, H-2'b), 2.64 (1H, m, H-3'a), 2.33 (1H, m, H-3'b), 2.30 (3H, s, H-4'), 2.30 (3H, s, H-5'), 3.28 (1H, m, H-6'a), 3.22 (1H, m, H-6'b), 1.16 (3H, t, J = 6.6 Hz, H-7'); 13C-NMR (CDCl3, 150 MHz) δC 32.4 (t, C-1), 26.2 (t, C-2), 76.7 (d, C-3), 40.4 (s, C-4), 42.6 (d, C-5), 18.0 (t, C-6), 32.0 (t, C-7), 40.8 (s, C-8), 54.1 (d, C-9), 36.7 (s, C-10), 126.8 (d, C-11), 125.5 (d, C-12), 135.9 (s, C-13), 41.5 (s, C-14), 24.2 (t, C-15), 30.4 (t, C-16), 52.0 (s, C-17), 134.1 (s, C-18), 40.7 (t, C-19), 33.0 (s, C-20), 37.7 (t, C-21), 29.7 (t, C-22), 71.3 (t, C-23), 17.4 (q, C-24), 18.0 (q, C-25), 16.7 (q, C-26), 19.5 (q, C-27), 178.2 (s, C-28), 24.7 (q, C-29), 31.9 (q, C-30), 45.5 (t, C-1'), 24.7 (t, C-2'), 55.2 (t, C-3'), 44.0 (q, C-4'), 44.0 (q, C-5'), 35.6 (t, C-6'), 14.6 (q, C-7'); ESIMS m/z 626 [M + H]+; HRESIMS m/z 626.4901 [M + H]+ (Calcd for C38H64O4N3, 626.4891).
3.5. Anhydride Coupling Esterification
To fatsicarpain A (2.0 mg) in CH2Cl2 (1.0 mL) at room temperature were successively added DMAP (1.0 mg), DMAP·HCl (0.1 mg) and EDC·HCl (1.0 mg) and the mixture was allowed to react overnight. The reaction was quenched by water, followed by extraction with EtOAc (3 × 1.5 mL). The EtOAc extract was successively washed with 5% aqueous HCl, saturated aqueous NaHCO3, and brine. The organic layer was dried over anhydrous MgSO4 and evaporated to give a crude product, which was subjected to a short silica gel column eluting with n-hexane–EtOAc (3:1) to yield 7 (1.7 mg). Similarly, 3α-hydroxyolean-11,13(18)-dien-28-oic acid (2.0 mg) and 3α-acetoxyolean-11,13(18)-dien-28-oic acid (5) (2.0 mg) in CH2Cl2 (1.0 mL) were submitted to anhydride coupling esterification with DMAP, DMAP·HCl and EDC·HCl at room temperature overnight to yield 8 (1.8 mg) and 9 (1.8 mg), respectively.
3α,23-Dihydroxyolean-11,13(18)-dien-28-oic anhydride (7). Yield 42%. white amorphous powder; [α]25D –24 (c 0.2, CHCl3); UV (MeOH) λmax (log ε) 237 (4.16), 248 (3.90), 264 (3.73) nm; IR (KBr) vmax 3,446, 2,937, 1,793, 1,772, 1,734, 1,457, 1,374, 1,243, 997 cm−1; 1H-NMR (CDCl3, 600 MHz) δH 1.41 (2H, m, H-1α and H-1'α), 1.69 (2H, m, H-1β and H-1'β), 1.53 (2H, m, H-2α and H-2'α), 2.01 (2H, m, H-2β and H-2'β), 3.73 (2H, br s, H-3 and H-3'), 1.76 (2H, br d, J = 10.8 Hz, H-5 and H-5'), 1.47 (4H, m, H-6 and H-6'), 1.43 (4H, m, H-7 and H-7'), 2.13 (2H, br s, H-9 and H-9'), 5.69 (2H, d, J = 10.2 Hz, H-11 and H-11'), 6.44 (2H, dd, J = 10.2, 2.4 Hz, H-12 and H-12'), 1.09 (2H, m, H-15α and H-15'α), 1.64 (2H, m, H-15β and H-15'β), 2.05 (2H, m, H-16α and H-16'α), 1.71 (2H, m, H-16β and H-16'β), 2.54 (2H, d, J = 14.4 Hz, H-19α and H-19'α), 1.80 (2H, dd, J = 14.4, 1.8 Hz, H-19β and H-19'β), 1.39 (2H, m, H-21α and H-21'α), 1.33 (2H, m, H-21β and H-21'β), 2.21 (2H, m, H-22α and H-22'α), 1.47 (2H, m, H-22β and H-22'β), 3.55 (2H, d, J = 11.4 Hz, H-23a and H-23'a), 3.43 (2H, d, J = 11.4 Hz, H-23b and H-23'b), 0.70 (6H, s, H-24 and H-24'), 0.94 (6H, s, H-25 and H-25'), 0.79 (6H, s, H-26 and H-26'), 1.00 (6H, s, H-27 and H-27'), 0.81 (6H, s, H-29 and H-29'), 0.96 (6H, s, H-30 and H-30'); 13C- NMR (CDCl3, 150 MHz) δC 32.5 (t, C-1 and C-1'), 26.3 (t, C-2 and C-2'), 77.0 (d, C-3 and C-3'), 40.4 (s, C-4 and C-4'), 42.6 (d, C-5 and C-5'), 18.0 (t, C-6 and C-6'), 31.9 (t, C-7 and C-7'), 40.8 (s, C-8 and C-8'), 54.2 (d, C-9 and C-9'), 36.7 (s, C-10 and C-10'), 127.4 (d, C-11 and C-11'), 125.1 (d, C-12 and C-12'), 137.6 (s, C-13 and C-13'), 42.2 (s, C-14 and C-14'), 24.9 (t, C-15 and C-15'), 31.7 (t, C-16 and C-16'), 49.8 (s, C-17 and C-17'), 130.2 (s, C-18 and C-18'), 40.0 (t, C-19 and C-19'), 32.4 (s, C-20 and C-20'), 36.8 (t, C-21 and C-21'), 35.4 (t, C-22 and C-22'), 71.3 (t, C-23 and C-23'), 17.4 (q, C-24 and C-24'), 18.0 (q, C-25 and C-25'), 16.6 (q, C-26 and C-26'), 19.8 (q, C-27 and C-27'), 172.7 (s, C-28 and C-28'), 24.2 (q, C-29 and C-29'), 32.1 (q, C-30 and C-30'); ESIMS m/z 945 [M+Na]+; HRESIMS m/z 945.6595 [M + Na]+ (calcd for C60H90O7Na, 945.6579).
3α-Hydroxyolean-11,13(18)-dien-28-oic anhydride (8). Yield 45%. white amorphous powder; [α]25D –84 (c 0.2, CHCl3); UV (MeOH) λmax (log ε) 236 (4.08), 248 (3.94), 264 (3.70) nm; IR (KBr) vmax 3,437, 2,937, 1,800, 1,770, 1,736, 1,457, 1,376, 1,243, 997 cm−1; 1H-NMR (CDCl3, 600 MHz) δH 1.38 (2H, m, H-1α and H-1'α), 1.63 (2H, m, H-1β and H-1'β), 1.60 (2H, m, H-2α and H-2'α), 2.02 (2H, m, H-2β and H-2'β), 3.44 (2H, br s, H-3 and H-3'), 1.35 (2H, m, H-5 and H-5'), 1.48 (4H, m, H-6 and H-6'), 1.33 (4H, m, H-7 and H-7'), 2.07 (2H, br s, H-9 and H-9'), 5.68 (2H, d, J = 10.8 Hz, H-11 and H-11'), 6.42 (2H, dd, J = 10.8, 3.0 Hz, H-12 and H-12'), 1.09 (2H, m, H-15α and H-15'α), 1.61 (2H, m, H-15β and H-15'β), 2.05 (2H, m, H-16α and H-16'α), 1.70 (2H, m, H-16β and H-16'β), 2.53 (2H, d, J = 14.4 Hz, H-19α and H-19'α), 1.78 (2H, d, J = 14.4 Hz, H-19β and H-19'β), 1.31 (2H, m, H-21α and H-21'α), 1.39 (2H, m, H-21β and H-21'β), 2.22 (2H, m, H-22α and H-22'α), 1.45 (2H, m, H-22β and H-22'β), 0.95 (6H, s, H-23 and H-23'), 0.84 (6H, s, H-24 and H-24'), 0.91 (6H, s, H-25 and H-25'), 0.77 (6H, s, H-26 and H-26'), 0.97 (6H, s, H-27 and H-27'), 0.80 (6H, s, H-29 and H-29'), 0.95 (6H, s, H-30 and H-30'); 13C-NMR (CDCl3, 150 MHz) δC 32.7 (t, C-1 and C-1'), 25.2 (t, C-2 and C-2'), 76.2 (d, C-3 and C-3'), 37.5 (s, C-4 and C-4'), 48.5 (d, C-5 and C-5'), 18.2 (t, C-6 and C-6'), 32.2 (t, C-7 and C-7'), 40.9 (s, C-8 and C-8'), 54.1 (d, C-9 and C-9'), 36.8 (s, C-10 and C-10'), 127.7 (d, C-11 and C-11'), 125.0 (d, C-12 and C-12'), 137.6 (s, C-13 and C-13'), 42.2 (s, C-14 and C-14'), 24.9 (t, C-15 and C-15'), 31.7 (t, C-16 and C-16'), 49.8 (s, C-17 and C-17'), 130.1 (s, C-18 and C-18'), 40.0 (t, C-19 and C-19'), 32.5 (s, C-20 and C-20'), 36.8 (t, C-21 and C-21'), 35.4 (t, C-22 and C-22'), 28.1 (q, C-23 and C-23'), 21.7 (q, C-24 and C-24'), 17.8 (q, C-25 and C-25'), 16.6 (q, C-26 and C-26'), 19.9 (q, C-27 and C-27'), 172.8 (s, C-28 and C-28'), 24.2 (q, C-29 and C-29'), 32.1 (q, C-30 and C-30'); ESIMS m/z 913 [M+Na]+; HRESIMS m/z 913.6683 [M + Na]+ (calcd for C60H90O5Na, 913.6680).
3α-Acetoxyolean-11,13(18)-dien-28-oic anhydride (9). Yield 47%. colorless needles; [α]25D –67 (c 0.2, CHCl3); UV (MeOH) λmax (log ε) 236 (4.18), 248 (3.98), 265 (3.75) nm; IR (KBr) vmax 2,935, 1,797, 1,771, 1,733, 1,456, 1,373, 1,241, 998 cm−1; 1H-NMR (CDCl3, 600 MHz) δH 1.26 (2H, m, H-1α and H-1'α), 1.65 (2H, m, H-1β and H-1'β), 1.66 (2H, m, H-2α and H-2'α), 1.96 (2H, m, H-2β and H-2'β), 4.66 (2H, br s, H-3 and H-3'), 1.29 (2H, m, H-5 and H-5'), 1.49 (4H, m, H-6 and H-6'), 1.35 (4H, m, H-7 and H-7'), 2.05 (2H, br s, H-9 and H-9'), 5.67 (2H, d, J = 10.2 Hz, H-11 and H-11'), 6.43 (2H, dd, J = 10.2, 2.4 Hz, H-12 and H-12'), 1.11 (2H, m, H-15α and H-15'α), 1.64 (2H, m, H-15β and H-15'β), 2.07 (2H, m, H-16α and H-16'α), 1.72 (2H, m, H-16β and H-16'β), 2.55 (2H, d, J = 15.6 Hz, H-19α and H-19'α), 1.79 (2H, d, J = 15.6 Hz, H-19β and H-19'β), 1.34 (2H, m, H-21α and H-21'α), 1.39 (2H, m, H-21β and H-21'β), 2.22 (2H, m, H-22α and H-22'α), 1.47 (2H, br d, J = 13.8 Hz, H-22β and H-22'β), 0.85 (6H, s, H-23 and H-23'), 0.89 (6H, s, H-24 and H-24'), 0.93 (6H, s, H-25 and H-25'), 0.78 (6H, s, H-26 and H-26'), 1.01 (6H, s, H-27 and H-27'), 0.81 (6H, s, H-29 and H-29'), 0.96 (6H, s, H-30 and H-30'), 2.09 (3H, s, 3-OAc); 13C NMR (CDCl3, 150 MHz) δC 33.3 (t, C-1 and C-1'), 22.7 (t, C-2 and C-2'), 78.3 (d, C-3 and C-3'), 36.7 (s, C-4 and C-4'), 49.8 (d, C-5 and C-5'), 18.0 (t, C-6 and C-6'), 32.1 (t, C-7 and C-7'), 40.9 (s, C-8 and C-8'), 54.1 (d, C-9 and C-9'), 36.7 (s, C-10 and C-10'), 127.4 (d, C-11 and C-11'), 125.1 (d, C-12 and C-12'), 137.6 (s, C-13 and C-13'), 42.2 (s, C-14 and C-14'), 24.9 (t, C-15 and C-15'), 31.7 (t, C-16 and C-16'), 49.8 (s, C-17 and C-17'), 130.2 (s, C-18 and C-18'), 40.0 (t, C-19 and C-19'), 32.5 (s, C-20 and C-20'), 36.8 (t, C-21 and C-21'), 35.4 (t, C-22 and C-22'), 27.7 (q, C-23 and C-23'), 21.3 (q, C-24 and C-24'), 17.8 (q, C-25 and C-25'), 16.6 (q, C-26 and C-26'), 19.8 (q, C-27 and C-27'), 172.7 (s, C-28 and C-28'), 24.2 (q, C-29 and C-29'), 32.1 (q, C-30 and C-30'), 170.8 (s, 3-OAc), 21.4 (q, 3-OAc); ESIMS m/z 997 [M + Na]+; HRESIMS m/z 997.6893 [M + Na]+ (calcd for C64H94O7Na, 997.6892). Crystal data: C64H94O7·H2O (formula weight 993.41), approximate crystal size, 0.64 × 0.56 × 0.30 mm, monoclinic, space group, P21, T = 110(2) K, a = 14.1986(4) Å, b = 13.3529(4) Å, c = 15.5987(5) Å, α = 90°, β = 104.518(3)°, γ = 90°, V = 2862.96(15) Å3, Dc = 1.152 Mg/m3, Z = 2, F(000) = 1088, μ(MoKα) = 0.074 mm−1. A total of 14049 reflections were collected in the range 2.93° < θ < 29.39°, with 9750 independent reflections [R(int) = 0.0216], completeness to θmax was 99.8%; psi-scan absorption correction applied; full-matrix least-squares refinement on F2, the number of data/restraints/parameters were 9750/1/681; goodness-of-fit on F2 = 1.046; final R indices [I > 2σ(I)], R1 = 0.0419, wR2 = 0.0808; R indices (all data), R1 = 0.0611, wR2 = 0.0838, absolute structure parameter −0.7(8), largest difference peak and hole, 0.423 and −0.206 e/Å3.