Chemical Constituents from Annona glabra III

Jour nal of the Chi nese Chem i cal So ci ety, 2000, 47, 913-920 913 Chemical Constituents from Annona glabra III Fang-Rong Chang ( ), Chung-Yi Chen ( ), Tian-Jye Hsieh ( ), Chun-Ping Cho ( ) and Yang-Chang Wu* ( ) Grad u ate In sti tute of Nat u ral Prod ucts, Kaohsiung Med i cal Uni ver sity, Kaohsiung 807, Tai wan, R.O.C. Twenty com pounds in clud ing a dioxoaporphine, annobraine (1); two oxoaporphines, liriodenine (2) and lysicamine (3); five aporphines, (-)-nornuciferine (4), (-)-anonaine (5), (-)-N-formylanonaine (6), (-)-asimilobine (7) and (+)-nordomesticine (8); one proaporphine, (+)-stepharine (9); two protoberberines, (-)-kikemanine (10) and dehydrocorydalmine (11); one azaanthraquinone, 1-aza-4-methyl-2- oxo-1,2- dihydro- 9,10-anthracenedione (12); two amides, N-trans-feruloyltyramine (13) and N-p-coumaroyltyramine (14); one ionone, blumenol A (15); and five ste roids, -sitosterol (16), stigmasterol (17), -sitosteryl-dglucoside (18), stigmasteryl-d-glucoside (19) and 6-O-palmitoyl- -sitosteryl-d-glucoside (20), were iso lated from the fruits and stems of Annona glabra. Among them, 1 is a novel dioxoaporphine al ka loid and 12 was ob - tained for the first time from nat u ral sources. These com pounds were char ac ter ized and ident i fied by phys i cal and spec tral ev i dence. IN TRO DUC TION struc ture char ac ter iza tion of com pounds 1 and 12. Annona glabra L. (Annonaceae), com monly known as pond-apple, is a trop i cal tree dis trib uted mainly in the Amer i - cas and south east Asia, and cul tured in the south ern part of Tai wan. It is used in tra di tional med i cines as an in sec ti cide and a parasiticide. 1,2 In pre vi ous stud ies, 3-22 we have in ves - ti gated the chem i cal con stit u ents of the Formosan Annonaceous plants, and have iden ti fied six new kaurane diterpenoids, annoglabasin A-F, along with 19 known kaurane diterpenoids from the fresh fruits and the stems of A. glabra. 10,22 Of these, methyl-16 -hy dro-19-al-ent- kauran- 17-oate ex hib ited mild ac tiv ity against HIV rep li ca tion in H9 lym pho cyte cells, and 16-17-dihydroxy-ent-kauran-19-oic acid showed sig nif i cant in hi bi tion of HIV-1 re verse tran scrip - tase. 10 Con tinuing in ves ti ga tion of these two parts of this plant led to the iso la tion of twenty pure sub stances, one isatinetype 23 dioxoaporphine, annobraine (1); two oxoaporphines, liriodenine (2) 17 and lysicamine (3); 17 five aporphines, (-)- nornuciferine (4), 17 (-)-anonaine (5), 17 (-)-N-formylanonaine (6), 6 (-)-asimilobine (7) 17 and (+)-nordomesticine (8); 24 one proaporphine, (+)-stepharine (9); 17 two protoberberines, (-)-kikemanine (10) 25 and dehydrocorydalmine (11); 26 one azaanthraquinone, 1-aza-4-methyl-2-oxo-1,2-dihydro-9,10- anthracenedi-one (12); 27 two amides, N-trans- feruloyl - tyramine (13) 17 and N-p-coumaroyltyramine (14); 11 one ionone, blumenol A (15); 28 and five ste roids, -sitosterol (16), 6 stigmasterol (17), 6 -sitosteryl-d-glucoside (18), 6 stigmasteryl-d-glucoside (19) 6 and 6-O-palmitoyl- - sitosteryl- D-glucoside (20). 29 The fol low ing de scribes the RE SULTS AND DIS CUS SION Annobraine (1) was ob tained as red nee dles from CHCl 3. HREIMS re vealed a [M] + ion at m/z 317.0690 (calcd 317.0688), cor re spond ing to the mo lec u lar for mula, C 19H 11NO 4. The EIMS re vealed frag ments at m/z 289 [M-28] + and 261 [M-56] +, which sug gested the ex is tence of two car - bonyl groups. The UV spec trum of 1 showed in tense ab sorp - tion bands at 232 (sh), 253, 280, 291, 333, 355 (sh) and 373 nm. The IR spec trum of 1 ex hib ited ab sorp tion bands at max 1727, 1036 and 968 cm -1 in di cat ing car bonyl and methyl - enedioxy groups, re spec tively. 17 1 H NMR spec trum of 1 ex - hib ited for four aryl pro ton sig nals at 8.57 (1H, dd, J = 7.6, 1.6 Hz), 7.62 (1H, td, J = 7.6, 1.6 Hz), 7.49 (1H, td, J = 7.6, 1.6 Hz), 8.83 (1H, dd, J = 7.6, 1.6 Hz), char ac ter is tic for H-8 ~ H-11, re spec tively, of oxoaporphines. A sin glet at 7.10 and a sin glet due to methylenedioxy pro tons at 6.35 were sim i lar to the A-ring sub sti tu tion of 2. 17 The 1 H NMR pat tern sug - gested the sub sti tu tion where the methylenedioxy group was placed at the 1,2-position and a sin glet at 7.10 was as signed to H-3. 17 The above spec tral data in di cated this com pound could be a gen eral oxoaporphine re lated to the known com - pound 2. 17, 25 But, two sig nif i cant two-proton trip lets at 3.95 (J = 6.4 Hz) and 3.31 (J = 6.4 Hz) in the aliphatic re gion over came this sup po si tion. Var i ous 2D NMR spec tra fur ther sup ported the struc ture of 1. Pro ton al lo ca tion of 1 was es tab lished by 1 H- 1 H COSY

914 J. Chin. Chem. Soc., Vol. 47, No. 4B, 2000 Chang et al. and 1 H- 1 H NOESY ex per i ments. Sig nif i cant cor re la tions be - tween H-3, H-4 and H-5, as well as H-8, H-9, H-10 and H-11, were ob served in the 1 H- 1 H NOESY spec trum (Fig. 1). Con - trasting with the 4,5-dioxoaporphine at 156.7 and 179.8 in the N-car bonyl and benzyl-carbonyl groups, 30 the sig nals of two car bonyl of 1 showed at 160.3 and 179.8. Three known isatine-type oxoaporphine, 31,32 telisatin A & B and laurodionine, showed char ac ter is tic chem i cal shifts by com - par i son with those of car bon yls of 1. No ob ser va tion of H-6a and H-7 sig nals in the 1 H NMR and the UV ab sorp tion sug - gested 1 as a 6a,7-dehydro aporphine with an ex tended con ju - ga tion, which in cluded a two-carbonyl bridge link be tween

Chem i cal Con stit u ents from Annona glabra III J. Chin. Chem. Soc., Vol. 47, No. 4B, 2000 915 N-6 and C-7. 31 13 C NMR data showed nine teen car bons, in - clud ing three meth y lene car bons at 27.5, 36.7 and 102.3, five methine car bons at 109.2, 123.4, 125.2, 127.5 and 129.1, and eleven qua ter nary car bons at 112.4, 119.8, 124.4, 126.7, 128.7, 129.6, 142.9, 151.5, 153.6, 160.3 (s, C-12), 179.8 (s, C-13), were con sis tent with the struc ture of 1. Due to the lim i ta tion of the sam ple amount, we failed to get its 2D 1 H- 13 C-correlation NMR spec tra. The as sign ments of 13 C NMR spec trum were di rectly com pared with known com - pounds, telisatin A & B, 31 and the re sults are de tailed in the com pound data of the ex per i men tal sec tion. Thus, 1 is a novel isatine-type 33 dioxoaporphine, which we named annobraine. Based on past semi-synthesis stud ies, 33,34 we pro posed that the biosynthetic hy poth e sis of isatine-type dioxoaporphine al ka - loids be pro duced by com bin ing N,6a- (or 6a,7-) dehydro - aporphine and ox alic acid through the en zy matic sys tem of this plant. Com pound 12, was iso lated as yel low nee dles. The UV and IR spec tral ab sorp tions sug gested that 12 should be sim i - lar to an anthraquinone ring sys tem. 17,27 Sig nif i cant peaks at m/z 239 [M] +, 211 [M-28] +, 183 [M-56] +, and 155 [M-84] + in the EIMS sys tem sup ported the three car bonyl groups, which re vealed 12 as an anthraquinone al ka loid by com par i son with data in the lit er a ture. 27 A methyl group at 2.64 (3H, d, J = 1.2 Hz) for C 4-CH 3 and 6.80 (1H, d, J = 1.2 Hz) for H-3 in the 1 H NMR of 12 are in agree ment with the pyridone moi ety. 17 The rel a tive sym me try of the chem i cal shift pat tern of the ar o matic pro tons with two-proton sig nals at 8.23 for pro tons peri to car bonyl group, and a multiplet (2H) at 7.72 could only be sat is fied by the par tial anthraquinoid-type struc ture 12. 17,29 The 13 C NMR spec trum of 12 showed four teen car bons, in - clud ing three iden ti cal car bonyl car bons at 162.2, 179.1 and 181.9. 35 The sub sti tu tions of 12 were con tem plated in an 1 H- 1 H NOESY ex per i ment. Sig nif i cant cor re la tions be tween the methyl group and H-3 as well as H-5, H-6, H-7 and H-8 were ob served in the 1 H- 1 H NOESY spec trum. The se quen tial cor - re la tions of the spec trum were suc cess fully es tab lished as shown in Fig. 2. The above re sults sup ported the struc ture of 12 as an 1-aza-4-methyl-2-oxo-1,2-dihydro-9,10-anthracene - dione. This com pound was pre pared syn thet i cally. 29,35 Com - par i son of the lit er a ture data 35 of nat u ral 12 with those of the syn thetic com pound proved the iden ti fi ca tion. Known com pounds were also iso lated and their struc - tures were es tab lished by spec tral meth ods. 6,17,24-29 EX PER I MEN TAL SEC TION General Methods UV spec tra were ob tained on a Hitachi 220-20 spectro - Fig. 1. NOESY Cor re la tions of 1. Fig. 2. NOESY Cor re la tions of 12.

916 J. Chin. Chem. Soc., Vol. 47, No. 4B, 2000 Chang et al. photometer in EtOH. IR spec tra were mea sured on a Hitachi 260-30 spectrophotometer. 1 H NMR (400 and 200 MHz) 13 C NMR NOESY and COSY spec tra were ob tained on a Varian NMR spec trom e ter. Low-resolution EIMS were re corded on a JEOL JMS-SX/SX 102A mass spec trom e ter or Quattro GC-MS spec trom e ter hav ing a di rect in let sys tem. High-resolution EIMS were mea sured on a JEOL JMS-HX 110 mass spec trom e ter. Sil ica gel 60 (Merck, 230-400 mesh) was used for col umn chro ma tog ra phy, precoated sil ica gel plates (Merck, Kieselgel 60 F-254, 0.20 mm) were used for an a lyt i cal TLC and precoated sil ica gel plates (Merck, Kieselgel 60 F-254, 0.50 mm) were used for pre para tive TLC. Spots were de tected by spray ing with Dragendorff s re agent or 50% H 2SO 4 and then heated on a hot plate. Plant Ma te rial The spec i men of A. glabra L. (Annonaceae) was col - lected from Pingtung County, Tai wan, Sep tem ber 1997. A voucher spec i men was iden ti fied by Dr. Hsin-Fu Yen and de - pos ited in the Grad u ate In sti tute of Nat u ral Prod ucts, Kaohsiung Med i cal Uni ver sity, Kaohsiung, Tai wan. Ex trac tion and Separation The fresh fruits (6.0 Kg) of A. glabra were ex tracted re - peat edly with MeOH (10 L 5) at room tem per a ture for 24-48 hrs. The com bined MeOH ex tracts were evap o rated and par ti - tioned to yield CHCl 3 and aque ous lay ers. The bases in the CHCl 3 so lu tion were ex tracted with 3% HCl to leave the acidic por tion and CHCl 3 so lu tion. The acidic por tion was basified with NH 4OH and ex tracted with CHCl 3. The CHCl 3 so lu tion was dried and evap o rated to leave a brown ish vis cous res i due (2.0 g). The res i due was fur ther sep a rated by col umn chro ma tog ra phy on Si gel with gra di ent sys tems of n-hex - ane-etoac (n-hex ane-etoac 4:1 to EtOAc) and EtOAc- MeOH (EtOAc to EtOAc-MeOH 4:1) to yield 60 frac tions of 120 ml each, which were fur ther com bined into 10 frac tions ac cord ing to their TLC pat terns. Each frac tion was chro - matographed over Si gel and pu ri fied by fur ther Si gel col umn chro ma tog ra phy, recrystalization, or pre para tive TLC to yield 6 com pounds. The yield amount and TLC data of these com - pounds are shown as fol lows: 2 (10.1 mg; CHCl 3-MeOH 9:0.5, R f 0.39), 4 (39.8 mg; CHCl 3 -MeOH 9:1, R f 0.35), 7 (27.9 mg; CHCl 3-MeOH 10:1, R f 0.29), 8 (4.4 mg; CHCl 3- MeOH 10:1, R f 0.64), 9 (5.0 mg; CHCl 3 -MeOH 9:1, R f 0.35) and 10 (6.7 mg; CHCl 3-MeOH 12:1, R f 0.30). The same pro ce - dures were also used on the stems. The air-dried stems (20.0 Kg) of A. glabra were ex tracted re peat edly with MeOH (10 L 6) at room tem per a ture for 24-48 hrs. The com bined MeOH ex tracts were evap o rated and par ti tioned to yield CHCl 3 and aque ous so lu tions. The bases in the CHCl 3 so lu tion were ex - tracted with 3% HCl to leave the acidic por tion (Part A) and CHCl 3 so lu tion (Part B, 20 g). The acidic por tion (Part A) was al ka lized with NH 4 OH and ex tracted with CHCl 3. The CHCl 3 so lu tion was dried and evap o rated to leave a brown ish vis cous res i due (3.5 g). The res i due was placed on a sil ica gel col umn chro ma tog ra phy and eluted with EtOAc grad u ally en riched with MeOH to af ford twenty frac tions. Each frac tion was rechromatographed over sil ica gel eluting with a gra di ent of CHCl 3/MeOH to ob tain 2 (5.0 mg, CHCl 3-MeOH 15:1, R f 0.45), 3 (3.3 mg; CHCl 3-MeOH 20:1, R f 0.48), 4 (15.0 mg; CHCl 3 -MeOH 10:1, R f 0.51), 5 (12.5 mg; CHCl 3 -MeOH 15:1, R f 0.42), 6 (3.4 mg; CHCl 3-MeOH 15:1, R f 0.40), 10 (23.1 mg; CHCl 3-MeOH 10:1, R f 0.25), and 11 (7.9 mg; CHCl 3-MeOH 6:1, R f 0.28), re spec tively. The CHCl 3 layer (Part B) was con - cen trated and chromatographed over sil ica gel us ing hex - ane/ac e tone as eluent to pro duce twenty-one frac tions. Each frac tion was rechromatographed over sil ica gel eluting with a gra di ent of n-hex ane/ac e tone to ob tain 1 (5.0 mg, CHCl 3 -MeOH 20:1, R f 0.66), 12 (4.2 mg; CHCl 3 -MeOH 10:1, R f 0.35), 13 (35.5 mg; CHCl 3-EtOAc 1:1, R f 0.50), 14 (8.2 mg, CH 2 Cl 2 -MeOH 10:1, R f 0.37), 15 (6.3 mg, CH 2 Cl 2 -MeOH 15:1, R f 0.54), mix ture of 16 and 17 (50.7 mg, CH 2Cl 2, R f 0.34), mix ture 18 and 19 (20.7 mg; CHCl 3-MeOH 10:1, R f 0.55), and 20 (11.1 mg; CHCl 3, R f 0.44), re spec tively. Annonbraine (1) Red nee dles (CHCl 3 ); mp 265-267 C; UV max/nm (EtOH) (log ) 232 (3.97, sh), 253 (4.37), 280 (3.80), 291 (3.84), 333 (3.64), 355 (3.43, sh) and 373 (3.20); IR max (neat) cm -1 1727, 1036 and 968; 1 H NMR (CDCl 3) /ppm 3.31 (2H, t, J = 6.4 Hz, H-4), 3.95 (1H, t, J = 6.4 Hz, H-5), 6.35 (2H, s, -OCH 2 O-), 7.10 (1H, s, H-3), 7.49 (1H, td, J = 7.6, 1.6 Hz, H-10), 7.62 (1H, td, J = 7.6, 1.6 Hz, H-9), 8.83 (1H, dd, J = 7.6, 1.6 Hz, H-11), 8.57 (1H, dd, J = 7.6, 1.6 Hz, H-8); 13 C NMR (CDCl 3) /ppm 27.5 (t, C-4), 36.7 (t, C-5), 102.3 (t, OCH 2O), 103.1 (s, C-7), 109.2 (d, C-3), 112.4 (s, C-1b), 123.4 (d, C-8), 124.4 (s, C-11a), 125.2 (d, C-10), 126.7 (s, C-7a), 127.5 (d, C-11), 128.7 (s, C-1a), 129.1 (d, C-9), 129.6 (s, C-3a), 142.9 (s, C-1), 151.5 (s, C-2), 153.6 (s, C-6a), 160.3 (s, C-12), 179.8 (s, C-13); EI-MS m/z (%) : 317 (100, M + ), 289 (66), 261 (43), 203 (37), 171 (17); HREIMS: m/z [M] + 317.0690 (calcd. for C 19 H 11 NO 4, 317.0688). Liriodenine (2) Yel low nee dles (CHCl 3 ); mp 280-282 C; UV max/nm (EtOH) (log ) 256 (4.21), 280 (3.98), 334 (3.92); IR max

Chem i cal Con stit u ents from Annona glabra III J. Chin. Chem. Soc., Vol. 47, No. 4B, 2000 917 (neat) cm -1 1650, 1050, 950; 1 H NMR (CDCl 3) /ppm 6.36 (2H, s, -OCH 2O-), 7.15 (1H, s, H-3), 7.56 (1H, td, J = 8.0, 0.7 Hz, H-9), 7.71 (1H, td, J = 8.0, 1.5 Hz, H-10), 7.74 (1H, d, J = 5.2 Hz, H-4), 8.57 (1H, dd, J = 8.0, 1.5 Hz, H-8), 8.60 (1H, dd, J = 8.0, 0.7 Hz, H-11), 8.87 (1H, d, J = 5.2 Hz, H-5); EI-MS m/z (%) 275 (100, M + ), 246 (18), 217 (7), 188 (39), 161 (16). Lysicamine (3) Yel low nee dles (CHCl 3); mp 185-187 C; UV max/nm (EtOH) (log ) 250 (4.60), 283 (4.50), 335 (3.82); IR max (neat) cm -1 1650; 1 H NMR (CDCl 3) /ppm 4.03 (3H, s, C 1- OCH 3), 4.11 (3H, s, C 2-OCH 3), 7.24 (1H, s, H-3), 7.58 (1H, td, J = 8.0, 1.1 Hz, H-9), 7.78 (1H, td, J = 8.0, 1.4 Hz, H-10), 7.82 (1H, d, J = 5.2 Hz, H-4), 8.59 (1H, dd, J = 8.0, 1.4 Hz, H-8), 8.93 (1H, d, J = 5.2 Hz, H-5), 9.19 (1H, dd, J = 8.0, 1.1 Hz, H-11); EI-MS m/z (%) 291 (99, M + ), 248 (100), 233 (17), 219 (5), 188 (14), 177 (30), 163 (9), 150 (16). (-)-Nornuciferine (4) Brown pow der (MeOH); mp 127-129 C; [ ] 24 D -58.8 (c = 0.18, CHCl 3 ); UV max/nm (EtOH) (log ) 230 (4.50), 272 (4.21), 310 (3.62); IR max (neat) cm -1 2900, 1590, 1440; 1 H NMR (CDCl 3 ) /ppm 2.72 (1H, t, J = 13.1 Hz, H-7 ), 2.88 (1H, dd, J = 13.1, 5.4 Hz, H-7 ), 2.93-3.15 (4H, m, H-4,5), 3.67 (3H, s, C 1-OCH 3), 3.46 (1H, dd, J = 13.1, 5.4 Hz, H-6a), 3.89 (3H, s, C 2 -OCH 3 ), 6.65 (1H, s, H-3), 7.20-7.33 (3H, m, H-8,9,10), 8.40 (1H, d, J = 7.6 Hz, H-11); EI-MS m/z (%) 281 (32, M + ), 266 (100), 250 (38), 238 (80), 223 (55), 206 (36), 194 (43). (-)-Anonaine (5) Pale yel low pow der (MeOH); mp 121-123 C; [ ] 24 D -53.0 (c = 0.54, CHCl 3); UV max/nm (EtOH) (log ) 230 (4.32), 272 (4.08), 310 (3.60); IR max (KBr) cm -1 1040, 950; 1 H NMR (CDCl 3) /ppm 2.64 (1H, t, J = 13.4 Hz, H-7 ), 2.85 (1H, dd, J = 13.4, 5.2 Hz, H-7 ), 2.73-3.38 (4H, m, H-4,5), 3.98 (1H, dd, J = 13.4, 5.2 Hz, H-6a), 5.93 and 6.08 (each 1H, d, J = 1.4 Hz, -OCH 2O-), 6.57 (1H, s, H-3), 7.23-7.26 (3H, m, H-8,9,10), 8.05 (1H, d, J = 7.4 Hz, H-11); EI-MS m/z (%) 265 (70, M + ), 264 (100), 248 (5), 236 (35), 235 (46). (-)-N-formylanonaine (6) (E-form : Z-form 2 : 1) Brown pow der (CHCl 3); mp 231-232 C; [ ] 24 D -106 (c = 0.2, CHCl 3 ); UV max/nm (EtOH) (log ) 223 (4.70), 268 (4.64), 290 (3.80), 316 (3.68); IR max (KBr) cm -1 2870, 1655, 1565, 1490, 1420, 1390, 1280, 1225, 1220, 1200, 1180, 1145, 1070, 1040, 935, 910, 850, 780, 740, 730, 640; 1 H NMR (CDCl 3) /ppm E-from: 2.74 (1H, dd, J = 10.8, 2.4 Hz, H-4 ), 2.84 (1H, dd, J = 14.0, 4.4 Hz, H-7 ), 2.89 (1H, m, H-4 ), 3.24 (1H, dd, J = 14.0, 4.4 Hz, H-7 ), 3.41 (1H, td, J = 12.0, 2.4 Hz, H-5 ), 3.82 (1H, ddd, J = 12.8, 4.8, 3.6 Hz, H-5 ), 5.06 (1H, dd, J = 13.6, 4.4 Hz, H-6a), 5.99 and 6.11 (each 1H, d, J = 1.2 Hz, -OCH 2O-), 6.59 (1H, s, H-3), 7.25-7.35 (3H, m, H- 8,9,10), 8.10 (1H, d, J = 7.6 Hz, H-11), 8.27 (1H, s, N=CHO - ); Z-from: 2.70-2.80 (2H, m, H-4,4 ), 2.90 (1H, dd, J = 14.0, 4.8 Hz, H-7 ), 3.11 (1H, m, H-5 ), 3.41 (1H, m, H-7 ), 4.36 (1H, dd, J = 13.6, 4.0 Hz, H-6a), 4.48 (1H, ddd, J = 12.8, 4.4, 3.6 Hz, H-5 ), 6.00 and 6.12 (each 1H, d, J = 1.2 Hz, -OCH 2O-), 6.62 (1H, s, H-3), 7.20-7.30 (3H, m, H-8,9,10), 8.12 (1H, d, J = 7.6 Hz, H-11), 8.40 (1H, s, N=CHO - ); EI-MS m/z (%) 293 (50, M + ), 263 (3), 262 (3), 249 (2), 248 (6), 237 (3), 236 (20), 235 (100), 205 (1), 204 (2), 179 (5), 178 (6), 177 (4), 176 (5), 152 (2), 151 (2). (-)-Asimilobine (7) Brown pow der (MeOH); mp 121-123 C; [ ] 24 D -49.0 (c = 0.56, CHCl 3); UV max/nm (EtOH) (log ) 222 (4.80), 274 (4.26), 308 (3.90); IR max (neat) cm -1 3500; 1 H NMR (CDCl 3) /ppm 2.69 (1H, t, J = 13.0 Hz, H-7 ), 2.86 (1H, dd, J = 13.0, 5.4 Hz, H-7 ), 2.90-3.00 (4H, m, H-4,5), 3.59 (3H, s, C 1-OCH 3), 3.83 (1H, dd, J = 13.0, 5.4 Hz, H-6a), 4.41 (1H, br s, C 2-OH), 6.69 (1H, s, H-3), 7.20-7.34 (3H, m, H-8,9,10), 8.29 (1H, d, J = 7.5 Hz, H-11); EI-MS m/z (%) 267 (78, M + ), 266 (100), 250 (38), 238 (80), 223 (55), 206 (36), 194 (43). (+)-Nordomesticine (8) Brown pow der (CHCl 3); mp 121-123 C; [ ] 24 D +17.1 (c = 0.07, CHCl 3 ); UV max/nm (EtOH) (log ) 216 (4.48), 282 (4.40), 309 (3.90); IR max (KBr) cm -1 3500, 1035, 965; 1 H NMR (CDCl 3) /ppm 3.90 (3H, s, C 2-OCH 3), 5.94 (1H, d, J = 1.4 Hz, -OCH 2 O-), 5.96 (1H, d, J = 1.4 Hz, -OCH 2 O-), 6.55 (1H, s, H-3), 6.72 (1H, s, H-8), 7.79 (1H, s, H-11); EI-MS m/z (%) 311 (59, M + ), 294 (26), 266 (19), 152 (18). (+)-Stepharine (9) Brown pow der (CHCl 3 ); mp 152-154 C; [ ] 24 D +110.0 (c = 0.10, CHCl 3); UV max/nm (EtOH) (log ) 217 (4.90), 233 (4.65), 290 (3.52); IR max (neat) cm -1 1705; 1 H NMR (CDCl 3 ) /ppm 3.60 (3H, s, C 1 -OCH 3 ), 3.81 (3H, s, C 2 - OCH 3), 6.29 (1H, dd, J = 10.0, 2.0 Hz, H-9), 6.40 (1H, dd, J = 10.0, 2.0 Hz, H-11), 6.65 (1H, s, H-3), 6.89 (1H, dd, J = 10.0, 2.8 Hz, H-8), 7.03 (1H, dd, J = 10.0, 2.8 Hz, H-12); EI-MS m/z (%) 297 (100, M + ), 296 (68), 282 (11), 268 (89), 253 (24), 237 (19), 225 (25), 209 (19).

918 J. Chin. Chem. Soc., Vol. 47, No. 4B, 2000 Chang et al. (-)-Kikemanine (10) Yel low pow der (CHCl 3); mp 83-85 C; [ ] 24 D -86.0 (c = 0.10, CHCl 3 ); UV max/nm (EtOH) (log ) 237 (4.04), 283 (3.78); IR max (neat) cm -1 3550, 2900, 1600; 1 H NMR (CDCl 3 ) /ppm 3.82 (3H, s, C 10 -OCH 3 ), 3.87 (3H, s, C 2 - OCH 3), 3.89 (3H, s, C 3-OCH 3), 6.62 (1H, s, H-1), 6.73 (1H, s, H-4), 6.80 (1H, d, J = 8.8 Hz, H-11), 6.84 (1H, d, J = 8.8 Hz, H-12); EI-MS m/z (%) 341 (53, M + ), 192 (100), 190 (49), 150 (29), 149 (38). Dehydrocorydalmine (11) Yel low pow der (CHCl 3); mp 85-88 C; UV max/nm (EtOH) (log ) 228 (4.90), 278 (4.74), 350 (4.10); IR max (neat) cm -1 3500, 2925, 1610; 1 H NMR (CDCl 3) /ppm 3.27 (2H, t, J = 6.4 Hz, H-5), 3.94 (3H, s, C 3-OCH 3), 3.99 (3H, s, C 2 -OCH 3 ), 4.15 (3H, s, C 9 -OCH 3 ), 4.92 (2H, t, J = 6.4 Hz, H-6), 7.05 (1H, s, H-4), 7.65 (1H, s, H-1), 7.78 (1H, d, J = 8.8 Hz, H-11), 7.90 (1H, d, J = 8.8 Hz, H-12), 8.73 (1H, s, H-13), 9.95 (1H, s, H-8); EI-MS m/z (%) 300 (5), 287(5), 259 (5), 241 (4), 187 (44), 148 (29), 147 (36). 1-Aza-4-methyl-2-oxo-1,2-dihydro-9,10-anthracenedione (12) 25 Yel low pow der; mp 249-251 C; [ ] D : 0 (c = 0.02, MeOH); UV max/nm (EtOH) (log ) : 258 (4.10), 294 (3.90); IR max (neat) cm -1 3026, 1710, 1664, 1598, 1523, 1477; 1 H NMR (C 5D 5N) /ppm 2.64 (3H, d, J = 1.2 Hz, C 4-CH 3), 6.80 (1H, d, J = 1.2 Hz, H-3), 7.72 (2H, m, H-6 and H-7), 8.23 (2H, m, H-5 and H-8); 13 C NMR (DMSO-D 6 ) /ppm 181.9 and 179.1 (CO-9 & CO-10), 162.2 (CO-2), 151.2 (C-3), 151.0 (C-4), 135.4 (C-6), 133.9 (C-7), 132.9 (C-10a), 130.7 (C-8a), 126.7 (C-8), 126.2 (C-5), 121.0 and 120.0 (C-4a and C-9a), 22.1 (CH 3); 35 EI-MS m/z (%) : 239 (100, M + ), 211 (55), 183 (24), 155 (27), 77 (69). N-trans-feruloyltyramine (13) Col or less crys tals (CHCl 3); mp 122-123 C; UV max/nm (EtOH) (log ) 220 (3.96), 293 (3.06), 319 (3.80); IR max (neat) cm -1 3300, 1650; 1 H NMR (CDCl 3 +CD 3 OD) /ppm feruloyl moi ety : 3.87 (3H, s, C 6-OCH 3), 6.15 (1H, d, J = 15.6 Hz, H-2), 6.83 (1H, d, J = 8.0 Hz, H-8), 7.00 (1H, dd, J = 8.0, 2.0 Hz, H-9), 6.95 (1H, d, J = 2.0 Hz, H-5), 7.46 (1H, d, J = 15.6 Hz, H-3); tyramine moi ety : 2.75 (2H, t, J = 6.8 Hz, H-2 ), 3.54 (2H, t, J = 6.8 Hz, H-1 ), 6.75 (2H, d, J = 8.8 Hz, H-5 and H-7 ), 7.02 (2H, d, J = 8.8 Hz, H-4 and H-8 ); EI-MS m/z (%) 313 (10, M + ), 192 (87), 177 (100), 145 (63), 120 (70), 107 (64). N-p-coumaroyltyramine (14) White pow der (CHCl 3); mp 235-238 C; UV max/nm (EtOH) (log ) 225 (4.12), 290 (3.50), 315 (4.16); IR max (neat) cm -1 3450, 1650, 1600, 1510; 1 H NMR (CD 3OD) /ppm coumaroyl moi ety : 6.36 (1H, d, J = 15.6 Hz, H-2), 6.79 (2H, d, J = 8.8 Hz, H-3 and H-5 ), 7.39 (2H, d, J = 8.8 Hz, H-2 and H-6 ), 7.43 (1H, d, J = 15.6 Hz, H-3); tyramine moi ety : 2.75 (2H, t, J = 7.2 Hz, H-2), 3.51 (2H, t, J = 7.2 Hz, H-1), 6.72 (2H, d, J = 8.8 Hz, H-3 and H-5 ), 7.02 (2H, d, J = 8.8 Hz, H-2 and H-6 ); EI-MS m/z (%) 283 (8, M + ), 164 (76), 147 (100), 120 (69), 107(90). Blumenol A (15) White pow der (CHCl 3); mp 114-116 C; [ ] 25 D +124.2 (c = 0.18, CHCl 3); IR (KBr) max 3400, 1760, 1670 cm -1 ; 1 H NMR (CDCl 3 ) /ppm 1.01 (3H, s, H-11), 1.07 (3H, s, H-12), 1.30 (3H, d, J = 6.4 Hz, H-10), 1.90 (3H, d, J = 1.4 Hz, H-13), 2.27 (1H, d, J = 17.1 Hz, H-2a), 2.49 (1H, d, J = 17.1 Hz, H-2b), 4.41 (1H, dq, J = 6.4, 6.4 Hz, H-8a), 5.80 (1H, d, J = 15.6 Hz, H-7), 5.90 (1H, dd, J = 15.6, 6.4 Hz, H-8b), 5.83 (1H, t, J = 2.5 Hz, H-4); 13 C NMR (CDCl 3 ) 41.2 (C-1, s), 49.7 (C-2, t), 198.0 (C-3, s), 126.8 (C-4, d), 162.8 (C-5, s), 79.0 (C-6, s), 129.0 (C-7, d), 135.8 (C-8, d), 68.0 (C-9, d), 18.9 (C-10, q), 24.0 (C-11, q), 23.7 (C-12, q), 22.9 (C-13, q); EIMS m/z (%) 224 ([M] +, 3), 206 (5), 168 (24), 150 (21), 135 (21), 125 (45), 124 (100). -Sitosterol (16) & Stigmasterol (17) White nee dles (MeOH); mp 138-140 C; [ ] 24 D -35 (c = 0.2, CHCl 3); IR max (neat) cm -1 3400, 2910, 1625, 1450; 1 H NMR (CDCl 3 ) /ppm 0.68 (3H, s, H-18), 0.81 (3H, d, J = 6.8 Hz, H-26), 0.84 (3H, d, J = 6.8 Hz, H-27), 0.86 (3H, t, J = 7.0 Hz, H-29), 0.92 (3H, d, J = 6.4 Hz, H-21), 1.01 (3H, s, H-19), 3.53 (1H, m, H-3), 5.02 (1H, dd, J = 16.1, 8.3 Hz, H-22), 5.12 (1H, dd, J = 16.1, 8.3 Hz, H-23), 5.36 (1H, br s, H-6); EI-MS m/z (%) 414 (80, M + ), 412 (30, M + ), 396 (45), 381 (27), 239 (33), 303 (33), 273 (23), 255 (33), 213 (29), 159 (25), 145 (29). -Sitosterol-D-glucoside (18) & Stigmasterol-D-glucoside (19) White pow der (MeOH); mp 198-200 C; [ ] 24 D -50 (c = 0.1, pyridine); IR max (KBr) cm -1 3400, 2910, 1625, 1450, 1360; 1 H NMR (CDCl 3 ) /ppm 0.68 (3H, s, H-18), 0.84 (3H, s, H-27), 0.86 (3H, t, J = 7.1 Hz, H-29), 0.94 (3H, d, J = 6.5 Hz, H-21), 1.01 (3H, s, H-19), 3.10-3-90 (5H, m, sugar moi ety H), 3.45 (1H, m, H-3), 4.35 (1H, d, J = 7.6 Hz, anomeric H), 5.02

Chem i cal Con stit u ents from Annona glabra III J. Chin. Chem. Soc., Vol. 47, No. 4B, 2000 919 (1H, dd, J = 12.3, 8.3 Hz, H-22), 5.12 (1H, dd, J = 12.3, 8.3 Hz, H-23), 5.25 (1H, d, J = 4.5 Hz, H-6); EI-MS m/z (%) 414 (100, M-glc), 397 (43), 396 (45), 382 (31), 354 (38), 329 (72), 303 (9), 275 (82), 255 (85). 6-O-Palmitoyl- -D-glucosyl- -sitosterol (20) Com pound 20 was pu ri fied by acetylation to yield tri - ace tate. The fol low ing is the phys i cal data of this tri ace tate. Tri ace tate: IR max (KBr) cm -1 1735, 1220, 1160, 1040, 900; 1 H NMR (CDCl 3) /ppm 0.67 (3H, s), 0.82 (3H, d, J = 5.6 Hz), 0.84 (3H, d, J = 6.1 Hz), 0.86 (3H, d, J = 6.1 Hz), 0.93 (3H, d, J = 6.1 Hz), 1.00 (3H, s), 1.26 (br s), 1.61 (2H, m), 2.01, 2.02, 2.04 (each 3H, s), 2.32 (2H, t, J = 7.3 Hz), 3.50 (1H, m), 3.65 (1H, m), 4.15 (1H, dd, J = 12.0, 2.8 Hz), 4.20 (1H, dd, J = 12.0, 5.0 Hz), 4.58 (1H, d, J = 8.0 Hz), 4.97 (1H, dd, J = 9.2, 8.0 Hz), 5.10, 5.17 (each 1H, t, J = 9.2 Hz), 5.36 (1H, d, J = 5.0 Hz). Hy dro lyzed tri ace tate of com pound 20 with NaOH in the mix - ture of MeOH and H 2 O to yield -sitosterol-d-glucoside (18) and fatty acid. The fatty acid was iden ti fied as palmitic acid by spec tral meth ods. By the EI-MS m/z (%) 284 (7, M + ), 85 (75), 71 (52), 57 (100). AC KNOWL EDG MENT This in ves ti ga tion was sup ported by a grant from the Na tional Sci ence Coun cil of the Re pub lic of China. We thank Pro fes sors Kuo-Hsiung Lee and Hui-Kang Wang (School of Phar macy, Uni ver sity of North Carolina at Cha pel Hill) for the mea sure ment of 13 C NMR spec trum of 1. Re ceived Feb ru ary 9, 2000. Key Words Annona glabra; Annonaceae; Al ka loids; 4-Methyl-1H-benzo<g> quinoline-2,5,10-trione; Annobraine. REF ER ENCES 1. Padmaja, V.; Thankamary, V.; Hara, N.; Fujimoto, Y.; Hisham, A. J. Ethnopharmacol. 1995, 48, 21. 2. Yang, T. H.; Chen, C. M. J. Chin. Chem. Soc.1995, 61, 146. 3. Chen, Y. Y.; Chang, F. R.; Wu, Y. C. J. Nat. Prod. 1996, 56, 904. 4. Chen, Y. Y.; Chang, F. R.; Yen, H. F.; Wu, Y. C. Phytochemistry 1996, 42, 1081. 5. Wu, Y. C.; Hung, Y. C.; Chang, F. R.; Cosentino, M.; Wang, H. K.; Lee, K. H. J. Nat. Prod. 1996, 59, 635. 6. Chen, C. Y.; Chang, F. R.; Wu, Y. C. J. Chin. Chem. Soc. 1997, 44, 313. 7. Chen, C. Y.; Chang, F. R.; Wu, Y. C. Tet ra he dron Lett. 1997, 38, 6247. 8. Chen, C. Y.; Chang, F. R.; Wu, Y. C. Tet ra he dron Lett. 1998, 39, 407. 9. Chang, F. R.; Chen, J. L.; Chen, Y. Y.; Chiu, F. F.; Wu, M. J.; Wu, Y. C. Phytochemistry 1998, 47, 1057. 10. Chang, F. R.; Yang, P. Y.; Lin, J. Y.; Lee, K. H.; Wu, Y. C. J. Nat. Prod. 1998, 61, 437. 11. Chen, C. Y.; Chang, F. R.; Yen, H. F.; Wu, Y. C. Phytochemistry 1998, 49, 1443. 12. Chang, F. R.; Wei, J. L.; Teng, C. M.; Wu, Y. C. Phytochemistry 1998, 49, 2015. 13. Chen, C. Y.; Wu, T. Y.; Chang, F. R.; Wu, Y. C. J. Chin. Chem. Soc. 1998, 45, 629. 14. Chang, F. R.; Wei, J. L.; Teng, C. M.; Wu, Y. C. J. Nat. Prod. 1998, 61, 1457. 15. Chen, C. Y.; Chang, F. R.; Chiu, F. F.; Wu, M. J.; Wu, Y. C. Phytochemistry 1999, 51, 429. 16. Chen, C. Y.; Chang, F. R.; Wu, Y. C. J. Chin. Chem. Soc. 1999, 46, 77. 17. Hsieh, T. J.; Chang, F. R.; Wu, Y. C. J. Chin. Chem. Soc. 1999, 46, 607. 18. Chang, F. R.; Chen, J. L.; Chiu, F. F.; Wu, M. J.; Wu, Y. C. Phytochemistry 1999, 51, 883. 19. Hsieh, T. J.; Chen, C. Y.; Kuo, R. Y.; Chang, F. R.; Wu, Y. C. J. Nat. Prod. 1999, 62, 1192. 20. Liaw, C. C.; Chang, F. R.; Wu, Y. C. J. Nat. Prod. 1999, 62, 1613. 21. Chang, F. R.; Chen, C. Y.; Wu, P. H.; Kuo, R. Y.; Chang, Y. C.; Wu, Y. C. J. Nat. Prod. 2000, 63, 746. 22. Chen, C. Y.; Chang, F. R.; Cho, C. P.; Wu, Y. C. J. Nat. Prod. 2000, 63, 1000. 23. Castedo, L.; Iglesias, T.; Puga, A.; Saá, J. M.; Suau, R. Heterocycles 1981, 15, 915. 24. Guinaudeau, H.; Leboeuf, M.; Cave, A. J. Nat. Prod. 1988, 51, 389. 25. Shama, M. The Isoquinoline Al ka loids; Ac a demic Press: New York, 1972; pp 269. 26. Suau, R.; Silva, M. V.; Valpuesta, M. Tet ra he dron 1991, 47, 5841. 27. Perez, J. M.; Vidal, L.; Grande, M. T.; Menendez, J. C.; Avendano, C. Tet ra he dron 1994, 50, 7923.

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