H-pyrazoles via room temperature Sonogashira arylation of propynal diethylacetal used as a propargyl aldehyde synthetic equivalent has been disclosed. The final acetal cleavage-cyclocondensation with hydrazine hydrochloride at 80 °C rapidly furnishes the title compounds in a one-pot fashion.A novel consecutive three-component synthesis of 3-(hetero)aryl-1 Theications ,5,6. In ications . In cropications . Furtherications ,10,11,12ications ,14, and ications ,16,17. Iications  and UV sications  as well ications ,21.Since multi-component reactions (MCRs) are highly efficient and address the fundamental principles of chemo-, regio-, and stereoselectivity, they are receiving an increasing interest in academia and industry ,33,34,35H-pyrazoles, i.e., monosubstituted pyrazole derivatives, via the concatenation of a Sonogashira alkynylation of (hetero)aryl iodides with the commercially available propynal diethylacetal, an in situ acetal cleavage, and a final cyclocondensation with hydrazine hydrochloride.In the past years, we have developed and elaborated the concept of MCR syntheses of multiple classes of heterocycles via Sonogashira coupling of alkynes and (hetero)aroyl chlorides with alkynyl ketones, followed by Michael addition-cyclocondensation, eventually in a one-pot fashion ,43,44,45Monosubstituted 3-(hetero)aryl pyrazoles can be retrosynthetically traced back either to (hetero)aroyl chlorides and TMS-acetylene via 3-trimethylsilylalk-2-yn-1-ones or to (hetero)aryl halides and propargyl aldehyde .Both disconnection approaches proceed via a retro alkynyl carbonyl condensation, thus leading to hydrazine and either to an alkynyl ketone ,47,48 orp-iodoanisole (1a) and propynal diethylacetal (2) into 1-(p-anisyl)-3,3-diethoxyprop-1-yne (3a) under standard Sonogashira conditions at room temperature was chosen as a model reaction .1,4-Dioxane is apparently a better solvent for the coupling than THF -3,3-diethoxyprop-1-yne (3a) in the presence of hydrazine hydrochloride [p-anisyl)-1H-pyrazole (4a)  , Table 2The optimal temperature of the sequential acetal cleavage-cyclocondensation is 80 °C , entry 1While the complete conversion is already achieved within 15 min , entry 5vide supra), the excess triethylamine from the coupling step has to be buffered (p-toluenesulfonic acid (PTSA)  , entry 41 in this novel consecutive three-component synthesis of 3-(hetero)aryl-1H-pyrazoles 4. After coupling of (hetero)aryl iodides 1 with propynal diethylacetal (2) at room temperature for two hours the subsequent addition of PTSA·H2O, hydrazine hydrochloride, and water, heating to 80 °C for 15 min gave rise to the isolation of 3-(hetero)aryl-1H-pyrazoles 4 in moderate to good yields aryl iodides d yields .This first study on the methodological scope of this novel three-component pyrazole synthesis shows that electron-rich as well as electron-poor aryl substituents can be readily introduced. Also, chloro and fluoro substituents and even unprotected phenol derivatives, which are incompatible with the coupling of acid chlorides, are well tolerated. Although a thienyl substituent can be carried through the sequence, all attempts to react other heteroaryl iodides such as 3- and 4-pyridyl iodides under the standard conditions met with failure.All cross coupling reactions were carried out in oven dried Schlenk or microwave tubes using septa and syringes under a nitrogen or argon atmosphere. Dry tetrahydrofuran and 1,4-dioxane were supplied by a MBraun system MB-SPS-800 solvent purification system. Chemicals were either commercially obtained from ABCR GmbH & Co KG, Acros Organics, Alfa Aesar GmbH & Co KG, Fluka, Merck KGaA, Riedel-de Haën, Sigma-Aldrich Co., and used as supplied or were already available in the research group.® 545 (0.02–0.10 mm) from Merck KGaA, Darmstadt before chromatographic purification. The reaction progress was observed qualitatively using TLC Silica gel 60 F254 aluminium sheets. The spots were detected with UV light at 254 nm and with aqueous potassium permanganate solution.All products were purified via column chromatography on silica gel 60 M (0.04–0.063 mm) from Macherey-Nagel using the flash technique under a pressure of 2 bar. The crude mixtures were absorbed on Celite1H-, 13C-, and 13C-135 DEPT NMR spectra were recorded on Bruker AVIII-300 spectrometer, using CDCl3 or DMSO-d6 as solvents. The resonances of CDCl3 or DMSO-d6 were locked as internal standards . The multiplicities of signals were abbreviated as follows: s: singlet; d: doublet; dd: doublet of doublets; dq: doublet of quartets; ddd: doublet of doublets of doublets; t: triplet; tdd: triplet of doublets of doublets; m: multiplet and br: broad signal. The type of carbon atom was determined on the basis of 13C-135 DEPT NMR spectra. For the description of the 13C-NMR spectra primary carbon atoms are abbreviated as CH3, secondary carbon atoms as CH2, tertiary carbon atoms as CH, and quaternary carbon atoms as Cquat. EI mass spectra were measured on Finnigan MAT 200 spectrometer. IR spectra were either obtained on Shimadzu IRAffinity or on Bruker Vector 22 FT-IR instruments. The intensity of the signals was abbreviated as follows: s (strong), m (medium), w (weak). The melting points (uncorrected) were measured on a Reichert Thermovar apparatus. Combustion analyses were carried out on a Perkin Elmer Series II Analyser 2400 in the microanalytical laboratory of the Institut für Pharmazeutische und Medizinische Chemie der Heinrich-Heine-Universität Düsseldorf.2(PPh3)2  and CuI  were tempered. Then, 1-iodo-4-methoxybenzene , 3,3-diethoxyprop-1-yne , degassed dry 1,4-dioxane (5.00 mL), and triethylamine  were successively added under an argon atmosphere. The mixture was stirred for 2 h until complete conversion of 1a (TLC monitoring). The crude mixture was absorbed onto Celite® and purified chromatographically on silica gel with hexanes/ethylacetate  to give the alkyne 3a  as a colorless oil. 1H-NMR : δ 1.27 , 3.65 , 3.80 , 3.81 , 5.48 , 6.80–6.85 , 7.38–7.43 . 13C-NMR : δ 15.1 (CH3), 55.2 (CH3), 60.8 (CH2), 83.0 (Cquat), 85.2 (Cquat), 91.7 (CH), 113.8 (CH), 113.9 (Cquat), 133.4 (CH), 159.9 (Cquat). EI + MS (m/z (%)): 234 , 205 ((M-C2H5)+, 2), 190 (16), 189 ((M-C2H5O)+, 100), 175 (12), 162 (20), 161 (95), 160 , 159 (22), 133 (28), 132 (18), 131 , 118 (12), 89 (15). IR : 2974 (w) cm−1, 2932 (w), 2884 (w), 2839 (w), 2224 (w), 1604 (w), 1570 (w), 1508 (s), 1458 (w), 1443 (w), 1391 (w), 1356 (w), 1329 (w), 1290 (m), 1246 (s), 1172 (m), 1090 (s), 1049 (s), 1032 (s), 1005 (s), 978 (m), 895 (w), 831 (s), 806 (m), 739 (w), 665 (w), 642 (w). Anal. calcd. for: C14H18O3 (234.3): C 71.77, H 7.74; Found: C 71.75, H 7.71.In a screw-cap Schlenk tube PdCl2(PPh3)2  and CuI  were tempered. Then, 1-iodo-3,4,5-trimethoxybenzene , 3,3-diethoxyprop-1-yne , degassed dry 1,4-dioxane (5.00 mL), and triethylamine  were successively added under an argon atmosphere. The mixture was stirred for 2 h at room temperature (water bath) until complete conversion of 1b (according to TLC monitoring). Then, hydrazine hydrochloride , deionized water (5.00 mL), and PTSA·H2O  were added. The mixture was stirred at 80 °C (preheated oil bath) until complete conversion (according to TLC monitoring). After extraction with CH2Cl2 (10 × 10 mL) the combined organic phases were dried with MgSO4. The solvents were removed in vacuo, the crude mixture was purified by chromatography on silica gel (CH2Cl2/MeOH/NH3 (aq)) to give 3--1H-pyrazole  as an orange solid, Mp 95 °C. 1H-NMR : δ 3.88 , 3.90 , 6.58 , 6.99 , 7.61 . 13C-NMR : δ 56.1 (CH3), 60.9 (CH3), 102.5 (CH), 103.0 (CH), 127.9 (Cquat), 132.6 (CH), 138.0 (Cquat), 149.5 (Cquat), 153.5 (Cquat). EI+MS (m/z (%)): 235 (13), 234 , 219 ((M−CH3)+, 68), 205 ((M−HN2)+, 2), 192 ((M−CH2N2)+, 2), 191 (30), 176 (30), 161 (20), 159 (15), 149 (13), 131 (15), 105 (22), 58 (23), 43 (67). IR : 3576 (w) cm−1, 3146 (w), 3041 (w), 2995 (w), 2963 (w), 2930 (m), 2860 (w), 2826 (w), 1730 (w), 1587 (m), 1516 (m), 1474 (m), 1449 (m), 1416 (m), 1325 (m), 1288 (w), 1261 (w), 1238 (m), 1121 (s), 1092 (m), 1057 (m), 1034 (w), 995 (s), 928 (w), 889 (w), 845 (s), 826 (m), 760 (s), 739 (m), 665 (w), 627 (m). Anal. calcd. for C12H14N2O3 (234.3): C 61.53, H 6.03, N 11.96; Found: C 61.63, H 6.29, N 11.68.In a screw-cap Schlenk tube PdCl4a, 4c–j were similarly prepared following the general procedure described above. The corresponding reactions conditions and work-ups are summarized in The pyrazoles 1H-NMR : δ 3.83 , 6.52 , 6.89–6.94 , 7.59 , 7.65–7.69 , 10.9 . 13C-NMR : δ 55.3 (CH3), 102.0 (CH), 114.1 (CH), 124.7 (Cquat), 127.1 (CH), 133.6 (CH), 148.5 (Cquat), 159.5 (Cquat). EI + MS (m/z (%)): 175 (12), 174 , 159 ((M−CH3)+, 52), 145 ((M−HN2)+, 2), 132 ((M−CH2N2)+, 5), 131 (37), 77 (11). IR : 3048 (w) cm−1, 2965 (w), 2914 (w), 2904 (w), 2824 (w), 2783 (w), 2725 (w), 1611 (w), 1526 (w), 1508 (m), 1454 (m), 1439 (m), 1417 (w), 1275 (m), 1248 (s), 1182 (s), 1113 (w), 1098 (m), 1055 (w), 1026 (s), 952 (m), 934 (m), 897 (m), 853 (m), 831 (s), 795 (m), 773 (s), 729 (w), 611 (m). Anal. calcd. for C10H10N2O (174.2): C 68.95, H 5.79, N 16.08; Found: C 68.73, H 5.48, N 15.88.195 mg  as a pale yellow solid. Mp 128 °C. 1H-NMR : δ 3.88 , 3.90 , 6.58 , 6.99 , 7.61 . 13C-NMR : δ 56.1 (CH3), 60.9 (CH3), 102.5 (CH), 103.0 (CH), 127.9 (Cquat), 132.6 (CH), 138.0 (Cquat), 149.5 (Cquat), 153.5 (Cquat). EI + MS (m/z (%)): 235 (13), 234 , 219 ((M-CH3)+, 68), 205 ((M-HN2)+, 2), 192 ((M-CH2N2)+, 2), 191 (30), 176 (30), 161 (20), 159 (15), 149 (13), 131 (15), 105 (22), 58 (23), 43 (67). IR : 3576 (w) cm−1, 3146 (w), 3041 (w), 2995 (w), 2963 (w), 2930 (m), 2860 (w), 2826 (w), 1730 (w), 1587 (m), 1516 (m), 1474 (m), 1449 (m), 1416 (m), 1325 (m), 1288 (w), 1261 (w), 1238 (m), 1121 (s), 1092 (m), 1057 (m), 1034 (w), 995 (s), 928 (w), 889 (w), 845 (s), 826 (m), 760 (s), 739 (m), 665 (w), 627 (m). Anal. calcd. for C12H14N2O3 (234.3): C 61.53, H 6.03, N 11.96; Found: C 61.63, H 6.29, N 11.68.295 mg  as an orange solid. Mp 60 °C. 1H-NMR : δ 6.73 , 6.93 , 7.05 , 7.24 , 7.61 , 7.64 , 9.9–11.2 . 13C-NMR : δ 102.1 (CH), 116.5 (Cquat), 117.0 (CH), 119.3 (CH), 126.6 (CH), 129.1 (CH), 129.3 (CH), 152.0 (Cquat), 155.8 (Cquat). EI + MS (m/z (%)): 161 (11), 160 , 132 (12), 131 ((M−HN2)+, 53), 104 (12). IR : 3277 (s) cm−1, 1736 (w), 1719 (w), 1686 (w), 1655 (w), 1625 (m), 1589 (s), 1560 (w), 1518 (m), 1451 (s), 1399 (m), 1286 (m), 1256 (s), 1200 (m), 1126 (m), 1110 (m), 1080 (m), 1047 (m), 952 (m), 826 (m), 779 (s), 746 (s), 600 (m), 561 (m), 517 (m). Anal. calcd. for C9H8N2O (160.2): C 67.49, H 5.03, N 17.49; Found: C 67.41, H 4.90, N 17.26.150 mg  as a yellow solid. Mp 92 °C. 1H-NMR : δ 6.60 , 6.68-6.70 , 7.15-7.31 , 7.7 , 9.4 , 12.4–13.5 .13C-NMR : δ 102.1 (CH), 112.2 (CH), 114.7 (CH), 116.3 (CH), 129.9 (CH), 135.3 (Cquat), 150.3 (Cquat), 157.8 (Cquat). EI + MS (m/z (%)): 161 (11), 160 , 131 ((M-HN2)+, 25), 93 . IR : 3245 (s) cm−1, 1610 (m), 1591 (s), 1541 (w), 1494 (m), 1478 (s), 1439 (s), 1382 (m), 1292 (m), 1258 (s), 1239 (m), 1218 (m), 1161 (w), 1117 (m), 1097 (m), 1082 (m), 1053 (m), 987 (w), 940 (m), 862 (s), 758 (s), 681 (s), 605 (w), 540 (w). Anal. calcd. for C9H8N2O (160.2): C 67.49, H 5.03, N 17.49; Found: C 67.50, H 5.04, N 17.31.196 mg  as a pale brown solid. Mp 154 °C. 1H-NMR : δ 6.56 , 7.07 , 7.59 , 7.69-7.74 , 11.0-12.2 . 13C-NMR : δ 102.5 (CH), 115.7 , 127.5 , 128.6 (Cquat), 132.5 (CH), 148.9 (Cquat), 162.6 . EI + MS (m/z (%)): 163 (11), 162 , 161 (11), 135 (16), 133 ((M-HN2)+, 27), 108 (11), 95 . IR : 3121 (w) cm−1, 3051 (w), 3034 (w), 2959 (w), 2918 (w), 2901 (w), 2882 (w), 2860 (w), 2841 (w), 2814 (w), 2776 (w), 2745 (w), 2725 (w), 2675 (w), 2627 (w), 2581 (w), 1651 (w), 1607 (w), 1524 (m), 1506 (m), 1452 (m), 1408 (w), 1352 (w), 1337 (w), 1233 (s), 1223 (s), 1204 (w), 1157 (m), 1111 (w), 1094 (m), 1082 (w), 1051 (m), 1015 (w), 953 (m), 934 (m), 885 (w), 839 (s), 812 (s), 770 (s), 752 (w), 692 (w), 673 (w), 602 (m). Anal. calcd. for C9H7FN2 (162.2): C 66.66, H 4.35, N 17.27; Found: C 66.92, H 4.41, N 17.00.170 mg  as a pale brown solid. Mp 104 °C. 1H-NMR : δ 6.60 , 7.36 , 7.60 , 7.69 , 8.5–12.1 . 13C-NMR : δ 102.8 (CH), 127.1 (CH), 128.9 (CH), 131.0 (Cquat), 132.2 (CH), 133.8 (Cquat), 149.0 (Cquat). EI + MS (m/z (%)): 180 (M(37Cl)+, 30), 179 (14), 178 (M(35Cl)+, 100), 151 ((M−HN2)(37Cl)+, 10), 149 ((M−HN2)(35Cl)+, 4), 116 (10), 115 (19), 113 , 111 , 89 (12). IR : 3165 (w) cm−1, 2955 (w), 2918 (m), 2876 (w), 2847 (w), 1510 (m), 1447 (m), 1406 (w), 1115 (w), 1090 (s), 1080 (m), 1047 (m), 1013 (m), 953 (m), 924 (m), 878 (w), 833 (s), 761 (s), 723 (m), 692 (m), 613 (m). Anal. calcd. for C9H7ClN2 (178.6): C 60.52, H 3.95, N 15.68; Found: C 60.78, H 4.15, N 15.55.121 mg  as an orange-brown solid. Mp 95 °C. 1H-NMR : δ 6.68 , 7.62 , 7.63 , 7.87 , 10.9–12.7 . 13C-NMR : δ 103.0 (CH), 124.1 , 125.7 , 125.9 (CH), 129.8 , 132.0 (CH), 135.9 (Cquat), 149.1 (Cquat). EI + MS (m/z (%)): 213 (11), 212 , 211 (12), 193 ((M−F)+, 7), 185 (11), 183 ((M−HN2)+, 5), 145 (14). IR : 3122 (w) cm−1, 3030 (w), 3013 (w), 2959 (w), 2918 (w), 2900 (w), 2868 (w), 2779 (w), 2727 (w), 2679 (w), 1620 (m), 1468 (w), 1416 (w), 1321 (s), 1275 (w), 1182 (s), 1159 (m), 1109 (s), 1097 (s), 1064 (s), 1053 (m), 1013 (m), 957 (m), 930 (w), 881 (w), 841 (s), 783 (m), 770 (s), 741 (m), 692 (m), 673 (w), 617 (w). Anal. calcd. for C10H7F3N2 (212.2): C 56.61, H 3.33, N 13.20; Found: C 56.83, H 3.08, N 13.04.244 mg  as a pale brown solid. Mp 162 °C. 1H-NMR : δ 6.70 , 7.66 , 7.68 , 7.89 , 10.4–12.5 . 13C-NMR : δ 103.5 (CH), 111.1 (Cquat), 118.9 (Cquat), 126.1 (CH), 131.3 (CH), 132.6 (CH), 137.2 (Cquat), 149.1 (Cquat). EI + MS (m/z (%)): 170 (12), 169 , 142 (15), 140 ((M−HN2)+, 12), 102 . IR : 3248 (w) cm−1, 3237 (w), 3129 (w), 3057 (w), 2980 (m), 2972 (m), 2889 (w), 2814 (w), 2220 (w), 1929 (w), 1609 (m), 1508 (w), 1464 (w), 1449 (w), 1393 (w), 1350 (w), 1233 (w), 1221 (w), 1184 (m), 1157 (w), 1074 (w), 1051 (m), 968 (w), 951 (s), 934 (w), 841 (s), 768 (s), 731 (m), 685 (w), 608 (s). Anal. calcd. for C10H7N3 (169.2): C 70.99, H 4.17, N 24.84; Found: C 70.78, H 4.10, N 24.59.90 mg  as a pale yellow solid. Mp 143 °C. 1H-NMR : δ 6.74 , 7.58 , 7.70 , 8.15 , 8.64 , 9.8–12.3 . 13C-NMR : δ 103.2 (CH), 120.6 (CH), 122.5 (CH), 129.7 (CH), 131.1 (CH), 131.6 (CH), 134.7 (Cquat), 148.7 (Cquat), 149.1 (Cquat). EI + MS (m/z (%)): 190 (12), 189 , 143 ((M−NO2)+, 33), 142 (10), 116 (39), 89 (28). IR : 3173 (w) cm−1, 3067 (w), 2961 (w), 2926 (w), 2851 (w), 1557 (w), 1530 (m), 1518 (m), 1504 (m), 1487 (m), 1398 (w), 1375 (m), 1344 (m), 1315 (w), 1275 (m), 1265 (w), 1221 (w), 1202 (w), 1072 (w), 993 (m), 972 (w), 897 (w), 851 (w), 829 (w), 775 (m), 735 (m), 698 (s), 683 (m), 671 (m), 615 (m). Anal. calcd. for C9H7N3O2 (189.2): C 57.14, H 3.73, N 22.21; Found: C 57.19, H 4.02, N 22.18.209 mg  as a pale brown solid. Mp 121 °C. 1H-NMR : δ 6.54 , 7.07 , 7.27 , 7.35 , 7.63 , 11.55 . 13C-NMR : δ 102.7 (CH), 124.2 (CH), 124.6 (CH), 127.6 (CH), 131.5 (CH), 135.7 (Cquat), 145.6 (Cquat). EI + MS (m/z (%)): 152 (13), 151 (26), 150 , 123 (20), 122 (16), 121 ((M−HN2)+, 35), 105 ((M−CHS)+, 7), 96 (22), 78 (12). IR : 3146 (w) cm−1, 3129 (w), 3117 (w), 3102 (w), 3022 (w), 2957 (w), 2913 (w), 2872 (w), 2855 (w), 2808 (w), 2361 (w), 1732 (w), 1560 (w), 1522 (w), 1464 (w), 1410 (w), 1371 (w), 1273 (w), 1179 (m), 1047 (m), 1030 (m), 910 (m), 845 (m), 824 (m), 756 (s), 723 (w), 692 (s), 644 (w), 602 (m). Anal. calcd. for C7H6N2S (150.2): C 55.97, H 4.03, N 18.65; Found: C 56.12, H 4.12, N 18.46.151 mg  as an orange solid. Mp 89 °C. H-pyrazoles 4 in moderate to good yields starting with the Sonogashira coupling of (hetero)aryl iodides 1 and the commercially available propynal diethylacetal (2) as a propargyl aldehyde synthetic equivalent, followed by a rapid sequential acetal cleavage-cyclocondensation with hydrazine hydrochloride. Further studies directed to the synthesis of luminescent pyrazole derivatives as ligands for metal organic frameworks (MOFs) are currently underway.In summary, we have disclosed a novel consecutive three-component synthesis of 3-(hetero)aryl-1


The -migration product endo2- was synthesized by efficient selective cyclopropanation of endo-5-vinylbicyclo[2.2.2]oct-2-ene at the exocyclic π-bond. Gas phase thermal reactions of exo1- afforded C8 epimerization to endo1-, - migrations to exo2- and endo2-, direct fragmentation to cyclohexadiene and vinylcyclopropane, and CPC rearrangement in the following relative kinetic order: epk > 13 k > fk > CPCk.The title compound  Conservation of Orbital Symmetry treatise, formally classified this type of skeletal rearrangement as a -sigmatropic carbon migration oct-2-enes are lower than those reported for bicyclo[3.2.0]hept-2-enes - sigmatropic migration including both si and sr products. This order is markedly different from the observation that -carbon shifts afford the dominant products in the corresponding bicyclo[3.2.0]hept-2-enes [Thermal reactions of bicyclo3.2.0]hept-2-enes and bicyclo4.2.0]oct-2-enes, despite their homologous relationship, afford different product stereoselectivities as well as preferred exit channels. The .2.0hept-.2.0oct-2t-2-enes , or metht-2-enes ,7. An eaexo-8-cyclopropylbicyclo[4.2.0]oct-2-ene (exo1-) can undergo -sigmatropic migration to the si (exo2-) and sr (endo2-) products or C8 epimerization to endo1- -to-homoallylic radical rearrangement of diradical A to diradical B  oct-2-ene (2). Our explanation for this high degree of regioselectivity is that the syn-hydrogens on the saturated -CH2CH2- bridge obstruct the exo face of the endocyclic olefin from reacting with the carbenoid complex. Similarly, the endo-vinyl group must also block the endo face of the endocyclic olefin. We thus observed relatively little endocyclic monocyclopropanation or dicyclopropanation, even when the reaction was allowed to proceed at room temperature. The reaction however ultimately reached a “steady state” when the ratio of endo2-:5 was 1.7:1. Preparative GC separation ultimately produced endo2- in greater than 98% purity by GC. Lewis acid-catalyzed Diels-Alder cycloaddition of 1,3-cyclohexadiene and acrolein with boron trifluoride yielded bicyclo[2.2.2]oct-5-en-2-carboxaldehyde  and exo1- (12.8 min), is suggestive of a bicyclic rather not a monocyclic structure. The potential bicyclic CPC product CPC-1 identified in m/z 79 has been ascribed to the 1,3-cyclohexadienyl cation that forms from the cyclohexenyl radical cation by loss of a hydrogen atom. Similarly, the pentenyl radical cation at m/z 68 can undergo loss of a hydrogen atom to form the 1,3-pentadienyl cation. More minor fragments correspond to a C7H10 radical cation at m/z 94 due to extrusion of 1,3-butadiene from the molecular ion and to a C9H12 radical cation at m/z 120 due to loss of ethylene from the molecular ion.The identity of a CPC product eluting at 12.3 min, a retention time intermediate between that of CPC-1 is based on independent synthesis using the synthetic route outlined in 6), a known compound, was prepared in high purity but low yield by Diels-Alder reaction of 1,3-cyclohexadiene and 2-chloroacrylonitrile followed by base-catalyzed hydrolysis  sigmatropic rearrangement and Cope rearrangement. Conversion of 8 to CPC-1 occurred through formation of the tosylhydrazone derivative of the ketone and subsequent Shapiro modification of the Bamford-Stevens reaction undeca-2,9-diene (10) as the sole thermal product, which eluted at 16.2 min using a standard GC program and, unlike the CPC thermal product that eluted at 12.3 min, exhibited a prominent mass spec fragment at m/z 94 due to a cycloheptadienyl radical cation. We assumed, given the comparability in carbon framework between compound 10 and bicyclo[5.4.0]undeca-3,8-diene, that 10 and bicyclo[5.4.0]undeca-3,8-diene would have similar GC retention times and mass spectral fragmentation patterns. An isomer of another potential CPC product, bicyclo[5.4.0]undeca-3,8-diene, was prepared using the synthetic methodology outlined in catalyst  proceedeexo1-, as shown in versus reaction time data for exo1- and for all isomeric products. As seen in endo2- < exo2- < CPC-1 < exo1- < endo1- < dodecane internal standard (ISTD). The value of the rate constant for overall loss of exo1-, 0k = 3.14 × 10−5 s−1, and its component rate constants were obtained using the Solver function in Microsoft Excel to fit experimental concentrations to the first-order exponential rate expressions based on the kinetic profile shown in ksi = 5.25 × 10−6 s−1, ksr = 2.92 × 10−6 s−1, epk = 1.9 × 10−5 s−1, k'f = 7.3 × 10−5 s−1, and kCPC = 6.5 × 10−7 s−1. The si/sr value thus derived from sik and srk is 1.8, and 13k = sik + srk = 8.2 × 10−6 s−1. The rate of direct fragmentation fk, as determined by curve fitting, is 2.9 × 10−6 s−1. The difference between 0k and the sum 13k + epk + kCPC approximated fk as 3.5 × 10−6 s−1. The relative order of importance of all kinetic processes is epk > 13k > fk >  CPCk  is 3.0, the highest among the four entries in 13k/fk ratio might represent a qualitative measure of the inward migratory aptitude of the migrating carbon.” We attribute the low contribution from fragmentation to steric interaction, which could be alleviated when C8 undergoes an endo trajectory, between the exo-cyclopropyl substituent at C8 and the syn-hydrogens at the other three cyclobutane carbons. While the absolute contribution from -carbon shifts is no greater for exo1- than for the exo-methoxy substrate (entry 3), the extent of epimerization is also greater for exo1-. It should be noted that the isomerization process that affords the epimeric product is also a likely outcome of inward migration. A rough measure of inward versus outward migration for C8 can be obtained by dividing the sum of the rate constants for formation of isomeric products (isomk = epk + 13k) by the rate constant for formation of the fragmentation product (fk), as seen in the next to last column in exo1- is ca. 10:1 for entry 4 compared to a value of ca. 3:1 for entry 3.According to Houk , “the dyt-2-enes . The exihanced 1,-migratioexo1-. The si/sr value of 1.8 for exo1- is consistent with a longer-lived transition structure that can undergo more extensive rotation before the migrating carbon reaches the migration terminus and collapses to form the si or sr products. The same stereochemical trends are also apparent for the bicyclo[3.2.0]hept-2-enes , si/sr ratio oct-2-en-6-one (3). Triethylamine , freshly distilled from CaH2, was dissolved in chloroform  and then added dropwise to a solution of cyclopropylacetyl chloride  in 1,3-cyclohexadiene . After stirring at rt for 24 h, the chloroform was removed via simple distillation. After addition of 150 mL of ether, the suspended solid was removed by vacuum filtration. The ether layer was washed with water and then brine, dried over MgSO4 (anhydrous), and concentrated under reduced pressure. After removal of hydrocarbon fractions on a silica gel column with pentane as the eluting solvent, the ketone product was eluted with 95:5 pentane:ether. Removal of solvent by rotary evaporation from five fractions yielded relatively pure ketone . 1H-NMR  δ 5.95 , 3.49 , 3.01 , 2.74 , 2.04 , 1.99 , 1.52 , 0.77 , 0.61 , 0.42 , 0.29 , 0.14 . 13C-NMR  δ 212.4 (C=O), 129.6 (=CH), 126.3 (=CH), 66.5 (CH), 54.5 (CH), 27.7 (CH), 21.3 (CH2), 18.4 (CH2), 6.5 (CH), 4.5 (CH2), 2.2 (CH2). FTIR (neat) νmax 3080, 3006, 1767, 1646, 685 cm−1. LRMS (EI) m/z 162 , 134 (5), 119 (5), 107 (8), 105 (10), 91 (32), 82 (27), 80 (100), 79 (61), 77 (25); HRMS (EI) calcd for C11H14O (M+) 162.1045, found 162.1043. 8-Cyclopropylbicyclo[4.2.0]oct-2-ene (1). A mixture of hydrazine sulfate  dissolved in hydrazine hydrate (10 mL) and compound 3  was refluxed overnight at 65 °C. The reaction mixture was extracted several times with ether, and the combined organic layers were washed with distilled water and brine, dried over MgSO4 (anhydrous), and concentrated under reduced pressure to afford crude hydrazone . FTIR (neat) νmax 3368, 3202, 3075, 3017, 1682, 1645, 1018, 739 cm−1. To a solution of potassium tert-butoxide , sublimed under high vacuum at 190 °C and dissolved in 25 mL anhydrous DMSO, was added 8-cyclopropylbicyclo[4.2.0]oct-2-en-7-one hydrazone  dropwise over 6 h. After stirring overnight, the reaction mixture was quenched with 5 mL of cold water and extracted four times with pentane. The combined pentane extracts were washed ten times with water to remove DMSO and dried over MgSO4 (anhydrous). The pentane was removed via simple distillation to yield crude product  in an exo:endo ratio of ca. 9:1. Preparative GC (on a 12' × ¼'' DC710 packed column at 130 °C) afforded exo1- in >99% purity by GC. 1H-NMR  δ 5.74 , 2.39 , 1.99 , 1.93 , 1.70 , 1.64 , 1.48 , 0.87 , 0.37 , 0.05 . 13C-NMR  δ 130.3 (=CH), 127.0 (=CH), 46.2 (CH), 38.4 (CH), 29.2 (CH), 28.2 (CH2), 25.7 (CH2), 22.3 (CH2), 15.5 (CH), 2.9 (CH2), 2.8 (CH2). FTIR (neat) νmax 3076, 3015, 1644, 1015, 697 cm−1. LRMS (EI) m/z 148 , 105 (3), 91 (8), 81 (9), 80 (100), 79 (45), 77 (11); HRMS (EI) calcd for C11H16 (M+) 148.1252, found 148.1255. Bicyclo[2.2.2]oct-2-en-5-carboxaldehyde (4). The catalyst BF3.OEt2  was added under inert atmosphere to a solution of 1,3-cyclohexadiene  and acrolein  dissolved in anhydrous ether (50 mL). After stirring for 4 h, the reaction was quenched with ice-cold distilled water and then extracted with ether. The combined organic layers were washed sequentially with water, saturated sodium bicarbonate (aq), water, and brine and then dried over MgSO4. Concentration via rotary evaporation resulted in crude product , 96% of the endo epimer by GC analysis. 1H-NMR  δ 9.44 , 6.32 , 6.10 , 2.95 , 2.63 , 2.55 , 1.72 , 1.65 , 1.55 , 1.35 , 1.27 . 13C-NMR  δ 203.6 (O=CH), 136.0 (=CH), 130.6 (=CH), 50.8 (CH), 30.6 (CH), 29.2 (CH), 26.6 (CH2), 25.1 (CH2), 24.7 (CH2). FTIR (neat) νmax 3044, 2937, 1721, 701 cm−1. LRMS (EI) m/z 136 , 108 (17), 93 (8), 91 (10), 80 (53), 79 (100).5-Vinylbicyclo[2.2.2]oct-2-ene (5). A solution of recrystallized methyltriphenylphosphonium bromide  in anhydrous THF (100 mL) was cooled to −78 °C and was treated slowly with 1.6 M CH3Li in ether . The resultant yellow suspension was then allowed to stir at 0 °C for 1 h. After again cooling the reaction mixture to −78 °C, bicyclo[2.2.2]oct-2-en-5-carboxaldehyde  dissolved in anhydrous THF (5 mL) was added dropwise via syringe over 1 h. The reaction mixture, which was allowed to warm gradually to rt and to stir overnight, was then quenched with ice-cold water and extracted twice with pentane. The combined organic layers were washed with water and brine, dried over MgSO4, and concentrated by short-path simple distillation to yield crude diolefin product  as a yellow oil. The endo:exo ratio, as determined by GC analysis, was 96:4. 1H-NMR  δ 6.28 , 6.14 , 5.55 , 4.86 , 4.79 , 2.46 , 1.75 , 1.55 , 1.46 , 1.24 , 1.06 . 13C-NMR  δ 145.4 (=CH), 134.8 (=CH), 132.0 (=CH), 111.6 (=CH2), 42.4 (CH), 35.7 (CH), 33.6 (CH2), 29.8 (CH), 26.1 (CH2), 24.4 (CH2). FTIR (neat) νmax 3040, 2934, 1638, 996, 906, 693 cm−1. LRMS (EI) m/z 134 , 92 (8), 91 (15), 80 (100), 79 (76); HRMS (EI) calcd for C10H14 (M+) 134.1096, found 134.1093. 5-Cyclopropylbicyclo[2.2.2]oct-2-ene (2). A solution of 1.0 M diethylzinc in hexanes  was added under argon to a dry round-bottomed flask at −40 °C. Sequential dropwise addition of 5-vinylbicyclo[2.2.2]oct-2-ene  and diiodomethane  was performed. The reaction mixture was then allowed to warm to −5 °C and the temperature was held between −5 °C and 0 °C for 16 h, during which time additional aliquots of diethylzinc (4.0 mL) and diiodomethane (0.2 mL) were added to facilitate selective cyclopropanation of 5-vinylbicyclo-[2.2.2]0ct-2-ene at the exocyclic vinyl group. After 16 h, the reaction vessel was sealed with parafilm and stored overnight in a freezer at 0 °C. GC analysis the following day showed a 1.0:1.7 ratio of 5-vinylbicyclo[2.2.2]oct-2-ene to 2, corresponding to a 63% conversion. The reaction was quenched with ice-cold water and extracted twice with pentane. The combined organic layers were washed with water and brine, dried over MgSO4, and concentrated by short-path simple distillation. The dominant epimer endo2- was obtained in >98% purity by prep GC (on a 12' × ¼'' DC710 packed column at 128 °C). 1H-NMR  δ 6.26 , 6.20 , 2.49 , 2.45 , 1.72 , 1.37 , 1.20 , 1.07 , 0.82 , 0.42 , 0.29 , 0.01 . 13C-NMR  δ 134.5 (=CH), 132.8 (=CH), 44.1 (CH), 35.3 (CH), 34.1 (CH2), 30.2 (CH), 26.4 (CH2), 24.6 (CH2), 18.1 (CH), 3.8 (CH2), 3.3 (CH2). FTIR (neat) νmax 3075, 3043, 3000,1640, 703 cm−1. LRMS (EI) m/z 148 , 92 (4), 91 (7), 80 (100), 79 (35); HRMS (EI) calcd for C11H16 (M+) 148.1252, found 148.1253. 2 at 20 °C resulted in 69% conversion. Selectivity for the desired product 2 was achieved regardless of temperature. The ratio of 2 to dicyclopropanated side product was 4.2:1 for the reaction at 20 °C compared to 5.0:1 for the reaction at 0 °C.A selective cyclopropanation of 5-vinylbicyclo[2.2.2]oct-2-ene to -endo2 was assessed by determining the -endo2:dodecane (ISTD) ratio over a period of 30 h at 275 °C; sampling at 10 h increments afforded a fundamentally invariant ratio of 5.2 ± 0.2. The thermal stability of -exo1 at 275 °C afforded a ca. 2:1 mixture of -endo2:-exo2. The LRMS of -exo2 was virtually indistinguishable from that of -endo2. The 13C-NMR signals due to -exo2 were identified from the NMR spectrum of this epimeric mixture: δ 136.2 (=CH), 133.7 (=CH), 42.5 (CH), 34.8 (CH), 32.6 (CH2), 30.2 (CH), 26.3 (CH2), 20.1 (CH2), 15.6 (CH), 4.0 (CH2).A preparative 120-h thermal reaction of Bicyclo[2.2.2]oct-5-en-2-one (6). Compound 6, a low melting solid prone to sublimation, was prepared as previously reported non-6-en-2-one (7). Subjecting 1.12 g (9.18 mmol) of 6 to Tiffaneau-Demjanov ring expansion [7 (contaminated with 7% bicyclo[3.2.2]non-6-en-3-one) after purification by column chromatography. 1H-NMR  δ 6.35 , 6.08 , 3.05 , 2.69 , 2.56 , 1.97 , 1.89 , 1.82 . 13C-NMR  δ 209.9 (C=O), 136.7 (=CH), 127.9 (=CH), 49.3 (CH), 39.4 (CH2), 31.2 (CH), 30.7 (CH2), 24.9 (CH2), 24.1 (CH2). FTIR (neat) νmax 3039, 2940, 1693, 705 cm−1. LRMS (EI) m/z 136 , 118 (50), 117 (38), 92 (85), 80 (57), 79 (100). xpansion  affordedBicyclo[5.2.2]undec-8-en-4-one (8). A sample of 0.34 g (2.5 mmol) 7 was allowed to react gradually with 4.0 mL (4.0 mmol) of 1.0 M vinylmagnesium bromide in THF under argon at −65 °C. Workup by treatment with 1 M NH4Cl (aq) followed by successive extraction with ether and subsequent drying over MgSO4 afforded 0.32 g  of crude vinyl alcohol as a pair of diastereomers. FTIR (neat) νmax 3424, 3032, 2975, 1631, 998, 916, 720 cm−1. LRMS (EI) m/z 164 , 146 (14), 131 (17), 104 (64), 92 (81), 91 (71), 83 (100), 79 (100). The vinyl alcohols were treated with 0.29 g (2.1 mmol) of KH  and 18-crown-6  dissolved in 20 mL anhydrous dioxane under reflux at 120 °C for 1.5 h. The major product in the crude mixture  corresponded to 8, as confirmed by 13C-NMR spectral comparison with reported literature values undeca-3,8-diene (CPC-1). To a solution of 150 mg (0.81 mmol) of p-toluenesulfonyl hydrazide in 1 mL of methanol was added crude 8 . The resultant crystals were filtered and washed with cold 1:1 pentane-ether (1 mL). The tosylhydrazone derivative of 8  was then suspended in TMEDA . After cooling the mixture in a dry ice/acetone bath, 1.6 M CH3Li in diethyl ether  was added dropwise via syringe. The resultant solution turned dark orange in color. After stirring overnight, the reaction was quenched with 1:1 pentane-water, washed successively with 2 M HCl (aq) and 2M NaOH (aq), and then dried over MgSO4. Removal of solvent by short-path distillation gave a trace amount of liquid. Major component LRMS (EI) m/z 148 , 133 (13), 120 (29), 106 (29), 93 (30), 92 (36), 91 (50), 79 (100). This mass spectrum is virtually identical to that of the thermal product eluting at 12.3 min (see SI).Bicyclo[5.4.0]undec-9-en-2-one (9). A sample of 1.63 g (12.2 mmol) of AlCl3 was weighed into a thick-walled flask under argon in a glovebag. A solution of 2-cycloheptenone  in anhydrous toluene (7.6 mL) was transferred to the flask via syringe. After complexation between the ketone and Lewis acid had proceeded for 40 min at rt, a solution of 1,3-butadiene in toluene  was added via syringe through a septum, which was then replaced with Teflon screw cap. After stirring the reaction at rt for 22 h, the reaction mixture was extracted with ether, and the organic layer was washed twice with water and once with brine. Elution of the ketone from a silica column with 98:2 pentane:ether afforded 1.04 g  of pure ketone 9. 1H-NMR  δ 5.68 , 5.62 , 2.76 , 2.65 , 2.45 , 2.29 , 2.17 , 1.89 , 1.80 , 1.66 . 13C-NMR  δ 215.2 (C=O), 125.4 (=CH), 125.3 (=CH), 49.7 (CH), 43.7 (CH2), 34.0 (CH), 33.3 (CH2), 31.7 (CH2), 27.5 (CH2), 26.2 (CH2), 24.1 (CH2). FTIR (neat) νmax 3023, 2921, 1691, 1654, 653 cm−1. LRMS (EI) m/z 164 , 146 (29), 135 (27), 117 (46), 104 (55), 91 (49), 79 (100), 77 (47). Bicyclo[5.4.0]undeca-2,9-diene (10). Ketone 9  was added to a solution of p-toluenesulfonyl hydrazide  in methanol (4 mL). After sitting overnight, a crop of crystals  was obtained. The tosylhydrazone of ketone 9 was dried in a vacuum oven: mp 137–139 °C. Dropwise addition of a solution of 1.6 M CH3Li in pentane  at −55 °C over 15 min to a suspension of the tosylhydrazone  suspended in 4 mL anhydrous TMEDA produced a dark red solution. After stirring overnight at rt, the reaction was quenched with cold water and extracted with pentane. Removal of the solvent by short-path distillation yielded product 10 , which was purified via prep GC (on a 12' × ¼'' DC710 packed column at 130 °C) for purposes of spectral analysis. 1H-NMR  δ 5.79 , 5.66 , 5.44 , 2.68 , 2.24 , 2.11 , 1.99 , 1.88 , 1.85 , 1.72 , 1.54 , 1.48 . 13C-NMR  δ 135.8 (=CH), 132.0 (=CH), 127.2 (=CH), 125.5 (=CH), 37.5 (CH), 35.3 (CH2), 34.6 (CH), 32.5 (CH2), 29.1 (CH2), 28.9 (CH2), 23.4 (CH2). LRMS (EI) m/z 148 , 133 (13), 119 (14), 105(21), 94 (84), 92 (31), 91 (48), 79 (100), 77 (25). si/sr = 1.8) observed for exo1- is consistent with a longer lifetime for diradical transition structure A  suggests a short lifetime for diradical transition structure B (CPC-1 is realized before intramolecular reorganization of the side chain results in other potential CPC products. The low stereoselectivity (ucture A  with moructure A . As confucture B  that has


Ring closure of β-amino ester 4 resulted in tricyclic pyrimidinones 15 and 16. The structures, stereochemistry and relative configurations of the synthesized compounds were determined by IR and NMR. all They reported that BCH, a non-metabolizable analog of leucine that mimics the stimulatory effect of glucose on insulin secretion, increased the incorporation of 3H-palmitic acid into protein heptane-2-carboxylic acid (BCH) blocks the transport of nonpolar amino acids across cell membranes, acts as an insulin-releasing factor and also inhibits the flavoprotein amino acid oxidases octane-2-carboxylic acid were prepared some years ago [-endo-3-aminobicyclo[2.2.2]oct-5-ene-2-carboxylic acid and its hydroxyl-substituted derivatives by stereoselective and regioselective functionalization of the double bond via 1,3-oxazine or γ-lactone intermediates. A further aim was a study of the ring-closure reactions of amino esters, and the retro-Diels-Alder reactions of the synthesized tricyclic pyrimidinones. ears ago ,21,22, b-endo-bicyclo[2.2.2]oct-5-ene-2,3-dicarboxylic acid anhydride (1) diastereoseletively. The starting di-endo-3-aminobicyclo[2.2.2]oct-5-ene-2-carboxylic acid (2) was prepared selectively by hypochlorite-mediated Hoffman degradation of the carboxamide obtained by ammonolysis of anhydride 1. Amino acid 2 was esterified in the presence of EtOH and SOCl2, furnishing the amino ester 4. Compound 2 was also transformed into cis-amino acid 3 with H2 in the presence of Pd/C, and it was protected with tert-butoxypyrocarbonate to give N-acylated amino acid 5  under two-phase conditions, furnishing iodolactone 6, which was reduced with Bu3SnH to give N-Boc lactone 7. When 7 was reacted with TFA or HCl, only the protecting group was eliminated, resulting in lactones 8a or 8b, instead of the all-endo-3-amino-6-hydroxybicyclo[2.2.2]octane-2-carboxylic acid. The similar lactone opening of 7 was also attempted with NaN3 octan-2-ol (10)  . N-acetyl derivative 11 was reacted with N-iodosuccinimide (NIS), a tricyclic dihydro-iodooxazine derivative was obtained regio- and stereoselectively. Not even traces of other regio- or diastereomers were observed in the crude product. Selective reduction of the halogen group with of this dihydro-iodooxazine Bu3SnH under an argon atmosphere led to the dihydrooxazine. Hydrolysis of this derivative with dilute HCl at room temperature gave N-acetylhydroxy amino acid 12. When 12 was boiled in acidic solution, all-endo-3-amino-5-hydroxybicyclo[2.2.2]octane-2-carboxylic acid hydrochloride (13) was produced in medium yield -one (15). In a similar manner as for the related tricyclic 3-substituted 2-thioxo-5,8-methanoquinazolin-4-ones investigated earlier, these compounds readily underwent decomposition when heated to their melting points; cyclopentadiene was split off, and monocyclic 2,3-dihydro-2-thioxopyrimidin-4(1H)-ones were formed oct-5-ene-2-carboxylic acid (2): di-endo-Bicyclo[2.2.2]oct-5-ene-2,3-dicarboxylic acid anhydride  was added in portions to dilute NH4OH  at 0 °C. The mixture was stirred for 30 min, and 2 M NaOH (60 mL) was then added dropwise at 0 °C over a period of 30 min, after which the excess of NH3 was removed under reduced pressure at 40 °C. The residue was cooled to 0 °C and 1 M NaClO solution (40 mL) was added dropwise with stirring, the temperature being maintained at 0 °C throughout. The mixture was stirred at the same temperature for 1 h, held at 70–75 °C for 10 min, then cooled to ambient temperature, adjusted with 10 M HCl to pH 7 and evaporated to dryness. The residue was extracted with three 150 mL portions of hot MeOH, and the extract was evaporated. The residue was dissolved in a small amount of water and the HCl was removed by means of a Dowex 50 ion-exchange column (acid cycle). Elution was effected with 1 M NH4OH solution. Each fraction was evaporated and the dry residue was dissolved in water, acetone was added until turbidity appeared, and the mixture was then allowed to stand in a refrigerator. The solid crystals were filtered off. Yield 3.35 g (66%); m.p. 204–208 °C C9H13NO2 (167.09): calcd. C 64.65, H 7.84; N 8.38, found C 64.77, H 7.96, N 8.43.cis-3-Aminobicyclo[2.2.2]octane-2-carboxylic acid (3): A solution of amino acid 2  and 10% Pd/C (100 mg) in MeOH (100 mL) was stirred under H2 (50 atm) for 3 days at room temperature. The Pd was then filtered off and the filtrate was concentrated under reduced pressure. The residue was crystallized from water-acetone. Yield 0.18 g (51%); a white solid, m.p. 215–220 °C, lit. m.p. 232–235 °C  oct-5-ene-2-carboxylate (4): SOCl2  was added dropwise with stirring to absolute EtOH (30 mL) at −10 °C. cis-3-Aminobicyclo[2.2.2]octane-2-carboxylic acid  was added in portions to the mixture, which was stirred for 30 min at 0 °C, and then for 3 h at room temperature, after which the mixture was refluxed for 1 h and next evaporated. The residue was crystallized from Et2O and recrystallized from EtOH/ Et2O). Yield 6.6 g (87%); a white solid, m.p. 209–213 °C, C11H18ClNO2 (231.10): calcd. C 57.02, H 7.83; Cl: 15.30, N 6.04, found C 57.22, H 7.92, Cl, 15.38, N 6.19.di-endo-3-tert-Butoxycarbonylaminobicyclo[2.2.2]oct-5-ene-2-carboxylic acid (5): 1 M NaOH (20 mL) was added to a solution of 3-aminobicyclo[2.2.2]oct-5-ene-2-carboxylic acid  in a 2:1 dioxane/H2O mixture (60 mL). The solution was cooled to 0 °C in an ice bath and di-tert-butyl dicarbonate  was added slowly. The mixture was stirred at 0 °C for 30 min and then warmed to room temperature and stirred for 4 h. The solvent was concentrated to 20 mL, the pH was then adjusted to 2.5 with 10% H2SO4, and the resulting solution was extracted with EtOAc (3 × 50 mL). The combined extracts were dried (Na2SO4) and evaporated, to give 5 as a white solid, which was recrystallized from iPr2O. Yield 3.4 g (63%); m.p. 117–120 °C C14H21NO4 (267.15): calcd. C 62.90, H 7.92, N 5.24, found C 62.78, H 7.99, N 5.11.3,7]decan-5-one-10-tert-Butoxycarbonylamino-2-iodo-4-oxatricyclodecan-5-one-10-tert-Butoxycarbonylamino-4-oxatricyclooctan-2-yl] carbamate (9): To a stirred suspension of LiAlH4  in dry THF (60 mL) was added a solution of Boc-lactone 7  in dry THF (20 mL). The resulting suspension was refluxed for 4 h and then decomposed by the addition of a mixture of water (2 mL) and THF (10 mL). The inorganic material was filtered off and washed with THF (3 × 50 mL). After drying (Na2SO4) and filtration, the solvent was evaporated off to give a pale oil, which was purified by column chromatography (toluene-MeOH = 4:1) Yield 0.41 g (81%) C14H25NO4 (271.18): calcd. C 61.97, H 9.29, N 5.16, found C 62.08, H 9.41, N 5.31.all-endo-5-Amino-6-(hydroxymethyl)bicyclo[2.2.2]octan-2-ol hydrochloride (10): Compound 9  was dissolved in aqueous HCl  and the solution was stirred at room temperature for 1 h. The solvent was next evaporated off and the residue was recrystallized from H2O-acetone. Yield 0.3 g, (72%); m.p. 165–167 °C. C9H18ClNO2 (207.10): calcd. C 52.05, H 8.74, Cl: 17.07, N 6.74, found C 52.24, H 8.92, Cl: 17.24, N 6.68.Ethyl di-endo-3-acetylaminobicyclo[2.2.2]oct-5-ene-2-carboxylate (11): To a suspension of ethyl 3-aminobicyclo[2.2.2]oct-5-ene-2-carboxylate hydrochloride  in CHCl3 (50 mL), Et3N , and AcCl  were added and the reaction mixture was stirred at room temperature for 2 h, and then washed with H2O (2 × 20 mL). The aqueous layer was extracted with EtOAc (3 × 30 mL). The combined organic layer was dried (Na2SO4) and evaporated. The residue was recrystallized from n-hexane-iPr2O. Yield 2.42 g (78%); m.p. 120–122 °C. C13H19NO3 (237.14): calcd. C 65.80, H 8.07, N 5.90, found C 65.94, H 8.32, N 5.78.all-endo-3-Acetylamino-5-hydroxybicyclo[2.2.2]octane-2-carboxylic acid (12): A solution of 11  in CH2Cl2 (80 mL) was treated with NIS  and subsequently stirred for 14 h at room temperature. When the reaction was completed, the mixture was washed with 10% NaOH solution (3 × 10 mL). The aqueous solution was extracted with CH2Cl2 (3 × 40 mL) and the organic phase was dried (Na2SO4) and evaporated. The oily dihydroiodooxazine product was sensitive to air and it was therefore used without purification in the next step.3SnH (4 mL) was added to a solution of oily dihydroiodooxazine (2.5 g) in dry CH2Cl2 (65 mL) under Ar. After stirring for 20 h at 40 °C, the solvent was evaporated off and the residue was purified by column chromatography on silica gel (n-hexane:EtOAc 10:1) to afford the dihydrooxazine derivative as a colorless oil . This oily product was also sensitive to air; it was therefore used immediately. A solution of oily dihydrooxazine derivative (1.05 g) in 20% aqueous HCl (20 mL) was stirred for 2 h. The solvent was then evaporated off to afford crude 12, which was recrystallized from H2O-acetone. Total yield 0.75 g (33%); m.p. 211–218 °C (with decomposition) C11H17NO4 (227.12): calcd. C 58.14, H 7.54, N 6.16, found C 58.26, H 7.72, N 6.32.Buall-endo-3-Amino-5-hydroxybicyclo[2.2.2]octane-2-carboxylic acid hydrochloride (13): A solution of 0.75 g (3.3 mmol) 12 in 20% aq. HCl (30 mL) was refluxed for 30 h. The solvent was then evaporated off to afford crude 9, which was recrystallized from EtOH-Et2O. Yield 0.5 g (67%); m.p. 222–230 °C (with decomposition) C9H16ClNO3 (221.08): calcd. C 48.76, H 7.27, Cl: 15.99, N 6.32, found C 48.64, H 7.12, Cl 16.14, N 6.38.Ethyl di-endo-3-phenylthiocarbamoylbicyclo[2.2.2]octane-2-carboxylate (14): To a magnetically stirred toluene solution of amino ester base 4 , one equivalent of PhNCS in toluene  was added dropwise . The mixture was refluxed for 10 h, the reaction mixture was then evaporated and the oily product was crystallized from n-hexane and recrystallized from iPr2O-EtOAc. Yield 0.68 g (57%); m.p. 110–112 °C. C18H22N2O2S (330.14): calcd. C 65.42, H 6.71, N 8.48 found C 65.61, H 6.59, N 8.58.-5,8-Ethano-3-phenyl-2-thioxo-2,3,4a,5,8,8a-hexahidroquinazolin-4(1H)-one (15): The thiocarbamoyl compound 14 (2.5 mmol) was refluxed in 20% aqueous HCl (30 mL) for 3 h. The reaction mixture was then evaporated, Et2O was added, and the crystalline product 15 was filtered off and recrystallized from iPr2O-EtOAc. Yield 0.68 g (57%); m.p. 285–289 °C. C16H16N2OS (284.10): calcd. C 67.58, H 5.67, N 9.85, found C 67.61, H 5.79, N 9.53.-2-(4-Chlorophenyl)-5,8-ethano-4a,5,8,8a-tetrahydroquinazolin-4(3H)-one (16): To a magnetically stirred toluene solution of amino ester base 4 , one equivalent of ethyl p-chlorobenzimidate in toluene  and a catalytic amount of p-toluene-sulfonic acid was added . The mixture was refluxed for 12 h, the reaction mixture was next evaporated and the residue was recrystallized from EtOH. Yield 0.65 g (63%); m.p. 210–215 °C. C16H15ClN2O (286.09): calcd. C 67.02, H 5.27, C: 12.36, N 9.77, found C 67.21, H 5.58, Cl: 12.48, N 9.68.Method A: -2-(4-Chlorophenyl)-5,8-ethano-4a,5,8,8a-tetrahydroquinazolin-4(3H)-one  was heated in a round-bottomed flask for 30 min at 220 °C. After the mixture had cooled, the residue was recrystallized from EtOH. Yield 0.12 g (58%).Method B: -2-(4-Chlorophenyl)-5,8-ethano-4a,5,8,8a-tetrahydroquinazolin-4(3H)-one  was refluxed in chlorobenzene (20 mL) for 12 h. The mixture was evaporated, and the residue was recrystallized from EtOH. Yield 0.13 g (63%). Method C: -2-(4-Chlorophenyl)-5,8-ethano-4a,5,8,8a-tetrahydroquinazolin-4(3H)-one  was weighed into a 10 mL pressurized reaction vial and the crystals were heated at 250 °C for 5 min at max. 300 W microwave irradiation. The crude product was recrystallized from EtOH. Yield 0.15 g (72%) m.p. 243–245 °C, lit. m.p. 245–246 °C, oct-5-ene-2-carboxylic acid derivatives, can be used for further valuable transformations, and are good starting materials for which the syntheses of hydroxy-substituted β-amino acids, aminodiols and heterocycles with potential biological activity.In summary, we have successfully synthetized di


In vitro antioxidant activity for 12 stannoxanes derived from Ph3SnCl (compounds 1-3), Ph2SnCl2 (compounds 4-6), Bu3SnCl (compounds 7-9), and Bu2SnCl2 (compounds 10-12), was assayed qualitatively by the chromatographic profile with 1,1-diphenyl-2-picrylhydrazil (DPPH) method and by two quantitative methods: the DPPH radical scavenging activity and Ferric-Reducing Antioxidant Power (FRAP) assays. The results were compared with those obtained with the starting materials 2-pyridine- carboxylic acid (I), 3-pyridinecarboxylic acid (II) and 4-pyridinecarboxylic acid (III), as well as with standard compounds, such as vitamin C and vitamin E, respectively. The in vitro antiradical activity with DPPH of diphenyltin derivative 5 showed a very similar behavior to vitamin C at a 20 μg/mL concentration, whereas according to the FRAP method, compound 8 was better. This difference is due to the mechanism of the antioxidant process. The Structure-Activity Relationships (SAR) for both methods is also reported. Bioorganometallic chemistry is dedicated to the study of metallic complexes as well as their biological applications, with a view to designing new drugs offering better performance than those already known. Molecules have been designed for the treatment of cancer ,2,3, AlzInvolved in organometallic chemistry, we are interested in the structural and pharmacological activity of metallic complexes of group 14. These elements have shown great chemical versatility, as a result of their diverse electronegativities and their ability to increase their coordination spheres . GermaniApplications of organotin(IV) compounds, especially those derived from carboxylate ligands, as bactericides ,14,15,162SnCl2) or triphenyltin chloride (Ph3SnCl) resulted in a higher antioxidant activity than that of kaempeferol or quercetin alone  Calc. C (61.06), H (4.6), N (2.97). Found C (61.01), H (4.12), N (2.84).Triphenyltin(IV) 3-pyridinecarboxylate (2). White powder, yield 24%, IR : 1426 (C=O sym), 1650 (C=O asym), 1592 (C=C py), 1479 (C=N py), 509 (Sn-C), 492 (Sn-O); NMR (CDCl3): 119Sn: ‑107; 1H: H2 (9.2), H4 (7.3), H5 (8.3), H6 (8.6), H9 (7.79), H10 (7.48), H11 (7.46); Elem. anal.: [C24H19NO2Sn] Calc. C (61.06), H (4.6), N (2.97). Found. C (60.66), H (4.05), N (2.90).Triphenyltin(IV) 4-pyridinecarboxylate (3). White powder, yield 86%, IR : 1344 (C=O sym), 1646 (C=O asym), 1543 (C=C py), 1473 (C=N py), 517 (Sn-C), 491 (Sn-O); NMR (CDCl3): 119Sn: -112; 1H: H2 (7.88), H3 (8.59), H5 (8.59), H6 (7.88), H9 (7.81), H10 (7.47), H11 (7.47); Elem. anal.: [C24H19NO2Sn] Calc. C (61.06), H (4.6), N (2.97). Found C (60.10), H (3.98), N (2.87).Diphenyltin(IV) 2-pyridinecarboxylate (4). White powder, yield 31%, IR : 1326 (C=O sym), 1673 (C=O asym), 1595 (C=C py), 1461 (C=N py), 526 (Sn-C), 473 (Sn-O); NMR (CDCl3): 119Sn: ‑427; 1H: H3 (8.35), H4 (7.98), H5 (7.51), H6 (8.67), H9 (7.52), H10 (7.2), H11 (7.19); Elem. anal.: [C24H18N2O4Sn∙C6H6] Calc. C (60.54), H (4.06), N (4.71); Found C (59.18), H (4.85), N (3.97). Diphenyltin(IV) 3-pyridinecarboxylate (5). White powder, yield 23%, IR : 1360 (C=O sym), 1651 (C=O asym), 1585 (C=C py), 1467 (C=N py), 491 (Sn-C), 490 (Sn-O); NMR (DMSO-d6): 119Sn: -233.5; 1H: H2 (9.07), H4 (7.56), H5 (8.37), H6 (8.71), H9 (7.85), H10 (7.26), H11 (7.26); Elem. anal.: [C24H18N2O4Sn∙C6H6] Calc. C (60.54), H (4.06), N (4.71); Found C (59.98), H (4.62), N (4.50).Diphenyltin(IV) 4-pyridinecarboxylate (6). White powder, yield 35%, IR : 1341 (C=O sym), 1641 (C=O asym), 1549 (C=C py), 1479 (C=N py), 534 (Sn-C), 492 (Sn-O); NMR (CDCl3): 119Sn: ‑233.5; 1H: H2 (7.58), H3 (8.33), H5 (8.33), H6 (7.58), H9 (7.73), H10 (7.14), H11 (7.14); Elem. anal.: [C24H18N2O4Sn∙C6H6] Calc. C (60.54), H (4.06), N (4.71); Found C (59.75), H (4.22), N (4.12).Tributyltin(IV) 2-pyridinecarboxylate (7). Yellow liquid, yield >90%, IR : 1395 (C=O sym), 1553 (C=O asym), 541 (Sn-C), 496 (Sn-O); NMR (CDCl3): 119Sn: +115.0; 1H: H3 (8.09), H4 (7.71), H5 (7.31), H6 (8.64), H8 (1.58), H9 (1.28), H10 (1.28), H11 (1.28); Elem. anal.: [C18H31NO2Sn] Calc. C (52.46), H (7.58), N (3.40). Found C (52.35), H (7.97), N (3.80).Tributyltin(IV) 3-pyridinecarboxylate (8). Yellow liquid, yield >90%, IR : 1344 (C=O sym), 1625 (C=O asym), 1460 (C=N py), 557 (Sn-C), 506 (Sn-O); NMR (CDCl3): 119Sn: +117.5; 1H: H2 (9.1), H4 (7.2), H5 (8.2), H6 (8.6), H8 (1.63), H9 (1.34), H10 (1.34), H11 (1.34); Elem. anal.: [C18H31NO2Sn] Calc. C (52.46), H (7.58), N (3.40). Found C (52.13), H (8.12), N (3.80).Tributyltin(IV) 4-pyridinecarboxylate (9). White powder, yield 23%, IR : 1348 (C=O sym), 1646 (C=O asym), 1603 (C=C py), 1460 (C=N py), 521 (Sn-C), 492 (Sn-O); NMR (CDCl3): 119Sn: +116.8; 1H: H2 (8.62), H3 (7.81), H5 (7.81), H6 (8.62), H8 (1.58), H9 (1.29), H10 (1.29), H11 (1.31); Elem. anal.: [C18H31NO2Sn] Calc. C (52.46), H (7.58), N (3.40). Found C (52.59), H (8.09), N (3.83).6H4NO2)(CH3O)Sn(C4H9)2]Dibutyltin(IV) methoxy-2-pyridinecarboxylate  Calc. C (46.67), H (6.53), N (3.63). Found C (45.91), H (7.06), N (3.91).6H4NO2)(CH3O)Sn(C4H9)2]Dibutyltin(IV)-methoxy-3-pyridinecarboxylate  Calc. C (46.67), H (6.53), N (3.63). Found C (45.41), H (7.15), N (3.83).6H4NO2)(CH3O)Sn(C4H9)2]Dibutyltin-methoxy-4-pyridinecarboxylate  Calc. C (46.67), H (6.53), N (3.63). Found C (45.41), H (7.33), N (3.99).et al. *100, where Ao corresponds to the absorbance of sample without stannoxane and Ae corresponds to the absorbance of sample with stannoxane.Antiradical activity evaluation for stannoxanes In this article, we decided to do an analysis of antiradical activity at 60 min (T = 60 min) at 20 μg/mL concentration in order to present the best activity for each compound and ligand and be able to evaluate the structure-activity relationships ,50. 1-12 as well as the ligands I-III to reduce iron(III) was carried out by the method reported by Hinneburg et al. [3Fe(CN)6, were added and this mixture was incubated for 30 min at 50 ºC, after this, 10% trichloroacetic acid solution (1000 μL) was added to the sample and this mixture was stirred for one minute and centrifuged at 1,500 rpm for 10 min; after the centrifugation a 1000 μL aliquot of the supernatant was taken and transferred to a clean assay tube where 0.1% FeCl3 solution (200 μL) was added. The absorbance of this mixture was recorded at 700 nm in an UV-spectrophotometer, the values are presented as means ±SD, n = 3.The ability of the stannoxanes g et al. , scaling5 > 6 = 11 in the DPPH quantitative assay and compounds 8 > 7 = 3 in the FRAP method. The most promising compound, in the context of this work, is compound 5. For DPPH, diphenyltin and dibutyltin (hexa- and pentacoordinated respectively) were the best, being the most active diphenyltin-bis-(3-pyridinecarboxylate) (compound 5) the most active. In the FRAP method tributyltin-3-pyridinecarboxylate  was the most active derivative. Two methods were used to determine the antioxidant power of stannoxanes; the proton and the electron donating ability were determined, and the results show that the bound and free pyridine and carboxylic nuclei were determinant for this, and the butyl- and phenyl groups play an important role in tin(IV) coordination, because of their electron-donating and electron-withdrawing capabilities. The results show that six of the twelve stannoxanes evaluated are good radical scavengers, compounds


Magnolia officinalis is a well-known Traditional Chinese Medicine. In the present study, two new alkaloids, named (S)-4-keto-magnoflorine (6) and (R)-3,4-dehydromagnocurarine (11), together with seven known alkaloids: (S)-magnoflorine (5), trans/cis N-feruloylputrescine (7/8), (R)-magnocurarine (10), (S)-tembetarine (12), (R)-oblongine (14), and (R)-asimilobine (17) were isolated and their structures elucidated by spectroscopic methods, including 1D, 2D NMR, and HRESI-MS. The absolute configurations of the isoquinoline alkaloids 5, 6, 10–12, 14, and 17 were determined by CD. In vitro inhibitory activities against aldose reductase, lipase, α-glucosidase, DPP–IV and three cancer cell lines  were evaluated for all isolated compounds. However, all compounds showed weak activities in all tests at the same concentration as the positive control drugs. An HPLC-ESI-MSn method has been established for screening of alkaloids in the bark of M. officinalis. A total of 23 alkaloids were identified or tentatively characterized; including 13 aporphines, eight benzylisoquinolines and two amides. Plausible fragmentation pathways of the representative compounds 6, 7/8, 11, and 17 were proposed in the present study.The bark of  Magnolia officinalis (Magnoliaceae), called Houpu in Chinese, has a very wide range of applications. It’s a common ingredient in the treatment of abdominal swelling of various causes and edema. Many of the prescriptions containing Houpu are also aimed at treatment of lung disorders (including cough and asthma) or intestinal disorders , -maine (17)  were isoα-glucosidase (40 µM), and DPP–IV (10 µM) were evaluated for all isolated compounds 5–8, 10–12, 14, and 17. All tested compounds showed weak activities at the same concentration as the positive control drugs . The inhibition rates of all tested compounds are all less than 20%. In the cytotoxicity bioassay, isolated compounds (5 µg/mL) showed weak activity against the A549, Bel-7402, and HCT-8 cancer cell lines, while the positive control 5-fluorouracil showed an IC50 value of 0.40 μg/mL. Inhibitory activities against aldose reductase (10 µM), lipase (10 µM), n spectra with those of the corresponding standards, peaks 5–8, 10–12, 14, and 17 could be unambiguously assigned as (S)-magnoflorine (5), (S)-4-keto-magnoflorine (6), trans/cis N-feruloylputrescine (7/8), (R)-magnocurarine (10), (R)-3,4-dehydromagnocurarine (11), (S)-tembetarine (12), (S)-oblongine (14), and (R)-asimilobine (17), respectively 2NH)]+ or [M+H–NH3]+ fragment ions as the base peak in their respective MS2 spectra, corresponding to quaternary and secondary aporphines, respectively + fragment ions in their MS2 spectra. Similarly, compounds 19, 20, and 22 should be secondary aporphines due to the significant [M+H–NH3]+ fragment ions in their MS2 spectra. The MS2 fragmentation of compounds 18, 21, and 23 provided significant [M+H–CH3NH2]+ ions in the product-ion spectra, suggesting one methyl group at the nitrogen atom of these compounds. The benzylisoquinoline alkaloids 10 and 12–16 showed obvious cleavages between C-1 and C-1a to produce the significant ion fragments at m/z 107 or 137, which were ascribed to hydroxybenzyl and 3-methoxy-4-hydroxybenzyl fragments, respectively + loss, whereas compounds 15 and 16 showed a characteristic [M−CH3NH2]+ loss indicating the number of methyl groups at the nitrogen atom. The ion peaks derived from loss of CH3OH, CH3O, H2O, CH3, CO and CH2O were characteristic of aporphine and benzylisoquinoline alkaloids, and served for their structural elucidation H]+ ions . Isolateectively . Based oectively . However3)2NH]+ fragment ion in the MS2 spectrum, but provided a significant and characteristic [M–C4H9N]+ fragment with m/z 285 as the base peak. Furthermore, the [M–C4H9N]+ fragment undergoes subsequent CH3OH, CO, OH, and CH3 losses. The plausible fragmentation pathway of compound 6 is shown in Isolated quaternary alkaloid 6 gave an abnormally weak 20 D +107 ; CD Δε (nm): +0.79 (346), –0.29 (297), +0.93 (235) ; 1H-NMR  δH: 6.87 , 6.55 , 6.40 , 3.91 , 3.75 , 3.70 , 3.31 , 2.82 , 2.78 , 2.16 ; 13C-NMR  δC: 189.4 (C-4), 165.2 (C-1), 154.3 (C-2), 151.5 (C-10), 146.6 (C-11), 134.3 (C-11c), 126.4 (C-7a), 124.1 (C-11b), 122.6 (C-11a), 121.2 (C-8), 116.3 (C-3a), 113.1 (C-9), 107.2 (C-3), 71.4 (C-6a), 69.9 (C-5), 58.1 (10-OMe), 58.0 (2-OMe), 56.2 (N-Me a), 48.0 (N-Me b); ESI-MS m/z: 356 [M]+; HRESI-MS m/z: 356.1514 [M]+ .(R)-3,4-Dehydromagnocurarine (11): White amorphous powder; UV (MeOH) λmax (nm): 227, 286; c, 0.17, MeOH); CD Δε (nm): +0.54 (297), –0.40 (264), –2.63 (230) ; 1H-NMR  δH: 7.07 , 6.91 , 6.71 , 6.68 , 6.21 , 5.90 , 4.60 , 3.87 , 3.52 , 3.43 , 3.22 , 2.92 ; 13C-NMR  δC: 157.4 (C-4'), 150.8 (C-6), 148.9 (C-7), 134.2 , 131.0 (C-3), 129.7 (C-1'), 127.6 (C-4), 125.8 (C-8a), 120.8 (C-4a), 119.3 (C-8), 118.0 , 114.4 (C-5), 76.2 (C-1), 58.9 (6-OMe), 55.1 (N-Me b), 53.3 (N-Me a), 35.7 (C-1a); ESI-MS m/z: 312 [M]+; HRESI-MS m/z: 312.1610 [M]+ .(S)-Magnoflorine (5): c, 0.15, MeOH); CD Δε (nm): +1.92 (317), –12.29 (267), +38.64 (233)  [(R)-Magnocurarine (10): c, 0.14, MeOH); CD Δε (nm): –1.14 (286), –9.59 (231) .(S)-Tembetarine (12): c, 0.11, MeOH); CD Δε (nm): +0.89 (286), +2.80 (237) . S)-oblongine (14): c, 0.23, MeOH); CD Δε (nm): –0.59 (272), +1.21 (234) .((R)-Asimilobine (17): c, 0.25, MeOH); CD Δε (nm): +11.04 (262), –79.13 (233) .See .See .See . 5–8, 10–12, 14, and 17 were isolated by the authors as described above. Their purities were above 95%, as determined by HPLC analysis.Compounds M. officinalis  was extracted with 25 mL 1% HCl in an ultrasonic bath (40 kHz) for 30 min. After cooling, the extracted solution was adjusted to the original weight and then filtered. An aliquot of 5 mL of filtrate was passed through Strata X-C strong cation resin (500 mg/6 mL). Wash the column with H2O (5 mL), MeOH (5 mL) and 2M NH4OH-MeOH (5 mL) successively. Collected the last elute and dry under the stream of N2. The residue resolved in MeOH (2 mL). After filtration with 0.22 μm filter, 10 μL of the aliquot was injected for HPLC–MSn analysis.Powdered stem bark of 3CN and (B) 0.1M ammonium acetate (adjust the pH to 7.5 using NH4OH). A gradient program was used as follows: 5–30% A at 0–20 min, 30–60% A at 20–35 min, and 60–90% A at 35–40 min. The column temperature was controlled at 35 °C. The flow rate was 1.0 mL/min. 283 nm was chosen as the detection wavelength. The optimized ESI-MSn parameters in the positive ion mode were as follows: skimmer, 40.0 V; capillary exit, 132.3 V; nebulizer, 42.0 psi; dry temperature, 350 °C; dry gas 12.0 L/min. For full-scan MS analysis, the spectra were recorded in the range of m/z 100–1000. Data-dependent scan program was used in the liquid chromatography/tandem mass spectrometry analysis so that the two most abundant ions in each scan were selected and subjected to MSn (n = 2–3) analyses. The collision-induced dissociation (CID) energy was adjusted to 45%. The isolation width of the precursor ions was 1.0 m/z.The sample was separated on a Gemini-NX C18 column . The mobile phase was (A) CHS)-4-keto-magnoflorine (6) and (R)-3,4-dehydromagnocurarine (11), were isolated from the stem bark of M. officinalis. A comprehensive study of alkaloid profile of M. officinalis has been conducted. Twenty three alkaloids including 13 aporphines, eight benzylisoquinolines and two amides were detected and tentatively identified by HPLC-MSn. Nine major alkaloids were unambiguously identified by comparison with isolated standards. In the present study, nine alkaloids including two new compounds, (


Their structures were elucidated by the spectral data as 26-O-β-D-glucopyranosyl-5α-furost-25 (27)-ene-3β, 12β, 22, 26-tetraol-3-O-β-D-glucopyranosyl (1→2) [β-D-glucopyranosyl (1→3)]-β-D-glucopyranosyl (1→4)-β-D-galactopyranoside (1) and 26-O-β-D-glucopyranosyl-5β-furost-20 (22)-25 (27)-dien-3β, 12β, 26-triol-3-O-β-D-glucopyranosyl (1→2)-β-D-galactopyranoside (2), respectively. Their cytotoxic activities on several cancer cell lines  were tested. 1 showed special cytotoxity on SF-268, while 2 showed cytotoxity on NCI-H460 and SF-268 cell lines, respectively.Two new steroidal saponins ( Allium macrostemon Bunge are well known as the traditional Chinese medicine “Xie bai”, which is used for treatment of throracic pain, stenocardia, heart asthma and diarrhea + . The 1H-NMR of 1 showed three methyl signals at δ 0.63 (s), 1.36 (s) and δ1.61 . Of all 57 carbon signals observed in 13C-NMR spectrum, 27 carbon signals , including a characteristic C25-C27 double bond carbon signals at δ 147.2 and 110.4. The 1H-NMR and 13C-NMR data indicated that 1 had the same sugar moiety as that of Macrostemonoside E -β-D-glucopyranosyl (1→4)-β-D-galactopyranoside.With the same NMR data of the sugar parts compared with Macrostemonoside E, the sugar moiety was finally unambiguously determined by acid hydrolysis and analysis of a combination of DEPT, 2 was also isolated as an amorphous powder. Its molecular formula of C45H72O19 was deduced by positive-ion HR-ESIMS at m/z 939.4572 [M+Na]+ . The 1H-NMR spectrum contained three steroid methyl groups at δ 0.92 , 0.99  and 2.00 . These three single peak methyl proton signals, especially the signal shift downfield to δ 2.00 compared with that of signal at H-21 of 1, revealed the presence of double bonds at C25-C27 and C20-C22. Comparison of the 13C-NMR spectrum of 2 with that of Macrostemonoside G + ; ESIMS (positive mode) at m/z 1,281 [M+ Na]+, 1,263 [M+ Na- H2O]+, 1,119 [M+ Na- 162]+, 1,101 [M+ Na- H2O- 162]+, 939 [M+ Na- H2O-162× 2]+, 777 [M+ Na- H2O- 162× 3]+; ESIMS (negative mode) at m/z 1,257 [M- H]-, 1,095 [M- H- 162]-, 933 [M-H-162×2]-, 771 [M- H- 162×3]-, 609 [M- H- 162× 4]-; IR νmax (KBr) cm -1: 3,420 (OH), 2,938 (CH), 1,000-1,100; 1H-NMR (C5D5N) and 13C-NMR data see 2Compound : An amorphous powder, mp 175-177°C; 2O, c 0.08); HR-ESIMS (positive mode) at m/z 939.4572 [M+Na]+ . FABMS (n= 1) at m/z 917 [M+ H]+, 755 [M+ H- 162]+, 737 [M+ H- 162- H2O]+, 593 [M+ H- 162× 2]+, 575 [M+ H- 162× 2- H2O]+, 431 [M+ H- 162×2- 162]+; IR νmax (KBr) cm -1: 3,410 (OH), 2,924 (CH), 1,010-1,120; 1H-NMR (C5D5N) and 13C-NMR data see 2OH·HCl (2 mg) were added to the residue, and the mixture was heated at 100 °C for 1 h. After cooling, Ac2O (0.5 mL) was added and the mixtures were heated at 100 °C for 1 h. The reaction mixtures were evaporated under reduced pressure, and the resulting aldononitrile peracetates were analyzed by GC-MS using standard aldononitrile peracetates as reference samples.Each saponin (5 mg) was heated in an ampoule with aq. 15% HCl (5 mL) at 110 °C for 2 h. The aglycon was extracted with dichloromethane three times and the aqueous residue was evaporated under reduced pressure. Then, pyridine (1 mL) and NHMCF-7, NCI-H460, SF-268 and HepG2 cells were maintained in RPMI 1640 (Gibco BRL) containing 10% FBS (Gibco), 2 mg/mL sodium bicarbonate, 100 µg/ml penicillin sodium salt and 100 µg/mL streptomycin sulfate. Cells were grown to 70% confluence, trypsinized with 0.05% trypsin-2 mM EDTA, and plated for experimental use. In all experiments, cells were grown in RPMI-1640 medium with 10% FBS for 24 hr prior to treatment.4 MCF-7, NCI-H460, SF-268 and HepG2 cells were seeded in 96 well tissue culture plates and treated with the two compounds on different concentration for 48 h. MTT  reagent  was added to each well and incubated for 4 h. After that, the suspended liquid was poured out and DMSO (100 μL) added to each well and swirled gently. Finally, the plate cover was removed and the absorbance in each well measured at 570 nm in a micro titer plate reader [1.0×10


O-methylnodulisporin F (1) and nodulisporin H (2), two new naphthoquinones, 5-hydroxy-2-methoxy-6,7-dimethyl-1,4-naphthoquinone (3) and 5-hydroxy-2-methoxynaphtho[9–c]furan-1,4-dione (4), and a new naphthofuran 1,3,8-trimethoxynaphtho[9–c]furan (5), along with five known compounds 4-O-methyl eleutherol (6), 2-acetyl-7-methoxybenzofuran (7), (-)-orthosporin (8), diaporthin (9), and 6-hydroxymellein (10), were obtained from the EtOAc extract of the mangrove-derived fungus Daldinia eschscholtzii HJ004. The structures of the isolated compounds were elucidated by extensive NMR and MS analyses, while the absolute configurations of the stereogenic carbons were established based on experimental and calculated electronic circular dichroism spectra. Compounds 4 and 7 displayed a potent inhibitory activity against α-glucosidase with the IC50 values of 5.7 and 1.1 μg/mL, respectively. Compounds 1 and 2 showed a moderate antibacterial activity against Staphylococcus aureus, methicillin-resistant S. aureus (MRSA) and Bacillus cereus, with minimum inhibitory concentration (MIC) values ranging from 6.25 to 12.5 μg/mL. Compound 3 exhibited antibacterial activity against B. cereus with the MIC value of 12.5 μg/mL.Two new polyketides, 8- In addition, in the sp3 region of the 1H NMR spectrum, with the help of DEPT-135 and HMQC spectra  and 3.98 , three methylenes at δH 3.12 , , 1.89 ), and 1.75 (m), and two methyls at δH 1.30  and 1.01  were observed, respectively. The 13C NMR spectrum showed 21 carbon signals, which, in combination with DEPT-135 and HSQC spectra , three methylene sp3 , two methine sp3 (δC 67.9 and 29.5), five methine sp2 , seven non-protonated sp2  and one carbonyl (δC 208.1) carbons. The above NMR data suggested that the structure of 1 was very similar to that of nodulisporin F -HR-ESI-MS at 247.0605 [M + H]+  (1H and 13C NMR data of 4 (3. The obvious difference was that Me-9 (δH 2.25/δC 11.9) and Me-10 (δH 2.36/δC 20.8) in 3 were replaced by two oxymethylene carbons at δC 72.1  and 74.3  in 4. Combination of the 1H and 13C NMR data with the molecular formula of 4 indicated the presence of a tetrahydrofuran ring. Therefore, the structure of 4 was established as 5-hydroxy-2-methoxynaphtho [9–c] furan-1,4-dione.Compound 13H11O5) . The 1H ata of 4  were sim5 was isolated as a yellow powder, with the molecular formula of C15H16O4 (eight degrees of unsaturation) based on [+]-HR-ESI-MS at m/z 261.1118 [M + H]+  , 6.71 , and 6.51 , three methoxyl signals at δH 3.97 , 3.89 , and 3.84 , and two oxymethylene signals at δH 5.26  and 5.16 . The 13C NMR spectrum , three protonated sp2 , two oxymethylene sp3 (δC 73.3 and 71.7) and three methoxyl  carbons. The HMBC correlations from H-9 (δH 5.16) to C-5 (δC 114.5), C-7 (δC 140.5) and C-10 (δC 71.7) as well as from H-10 (δH 5.26) to C-6 (δC 127.7) and C-8 (δC 150.6) suggested the presence of a naphthofuran skeleton. Furthermore, the HMBC correlations from OMe-1(δH 3.97) to C-1 (δC 157.5), OMe-3 (δH 3.89) to C-3 (δC 158.1), and OMe-8 (δH 3.84) to C-8 (δC 150.6) established the positions of the three methoxyls at C-1, C-3 and C-8, respectively. Thus, the structure of 5 was elucidated as 1,3,8-trimethoxynaphtho [9–c] furan.Compound 15H17O4) . The 1H NMR data , assigneNMR data , exhibitspectrum , in combspectrum , display4 and 7 displayed potent inhibitory activity on α-glucosidase with the IC50 values of 5.7 and 1.1 μg/mL, respectively. The rest of the isolated compounds exhibited no inhibitory activity against α-glucosidase. Acarbose was used as a positive control (IC50 = 2.0 μg/mL).Compounds 1 and 2 showed moderate antibacterial activity against Staphylococcus aureus, Methicillin-resistant S. aureus MRSA and Bacillus cereus, with minimum inhibitory concentration (MIC) values ranging from 6.25 to 12.5 μg/mL  instrument by using CDClterature .Brguiera sexangula var. rhynchopetala, collected in the South China Sea, and was identified as Daldinia eschscholtzii with the GeneBank (NCBI) accession number MH059553 25D = −17.8° ; UV (MeOH) λmax (log ε) 210 (2.31), 228 (2.42); IR (KBr) νmax 3261, 1636, 1270 cm−1; CD  λmax (Δε) 212 (+0.6), 239 (−1.2) nm. 1H and 13C NMR data see m/z: 357.1693 [M + H]+ .Nodulisporin H (2): white amorphous powder; [α]25D = +10.6° ; UV (MeOH) λmax (log ε) 217 (3.21), 260 (2.12); IR (KBr) νmax 3259, 1620, 1256 cm−1; CD  λmax (Δε) 216 (−3.0), 256 (−0.7) nm. 1H and 13C NMR data see m/z: 357.1693 [M + H]+ .5-Hydroxy-2-methoxy-6,7-dimethyl-1,4-naphthoquinone (3): yellow powder; UV (MeOH) λmax 282 (4.20), 249 (3.40), 217 (3.26) nm; IR (KBr) νmax 3240, 1865, 1680 cm−1; 1H and 13C NMR data see m/z: 233.0809 [M + H]+ .5-Hydroxy-2-methoxynaphtho [9–c] furan-1,4-dione (4): brown powder; UV (MeOH) λmax 301 (3.80), 250 (3.12), 217 (2.90) nm; IR (KBr) νmax 3246, 1864, 1680 cm−1; 1H and 13C NMR data see m/z: 247.0605 [M + H]+ .1,3,8-Trimethoxynaphtho [9–c] furan (5): yellow powder; UV (MeOH) λmax 278 (3.60) nm; IR (KBr) νmax 3500, 1632 cm−1; 1H and 13C NMR data see m/z: 261.1118 [M + H]+ .α-glucosidase inhibitory activity of 1–10 was determined using the procedure reported by Sawada et al.  furan-1,4-dione (4), and a new naphthofuran 1,3,8-trimethoxynaphtho [9–c] furan (5), and five known compounds (6–10) were obtained. Compounds 4 and 7 displayed potent inhibitory activity against α-glucosidase with the IC50 values of 5.7 and 1.1 μg/mL, respectively. Compounds 1 and 2 showed moderate antibacterial activity against S. aureus, Methicillin-resistant S. aureus MRSA and B. cereus with MIC values ranging from 6.25 to 12.5 μg/mL. Compound 3 exhibited antibacterial activity against B. cereus with the MIC value of 12.5 μg/mL.From the mangrove-derived fungus


OUCMDZ-302 with the mangrove plant, Excoecaria agallocha (Euphorbiaceae). The new structures of 2–6 were established on the basis of ECD, specific rotation and spectroscopic data as well as the chemical calculation. Compound 7 showed cytotoxicity against H1975 cell line with an IC50 value of 10.0 μM. Compounds 4 and 8–10 showed radical scavenging activity against DPPH with the IC50 values of 2.65, 0.24, 5.66, and 6.67 μM, respectively. In addition, the absolute configuration of compound 1 was solidly determined by X-ray and sugar analysis of the acidic hydrolysates for the first time as well as those of compounds 8–10 in this paper.Five new polyketides ( Excoecaria agallocha (Euphorbiaceae), also known as blind-your-eye, is mainly used to treat sores and stings. More than 72 cytotoxic diterpenoids have been identified from E. agallocha, structurally belonging to labdane  -7-O-α-D-ribofuranosyl-5-hydroxy-2-methylchroman-4-one (1)   -4-methoxy-2-methylchroman-5-ol (9) -2-methylchroman-4,5-diol (10)       Figure  isolateds Figure , (2R)-7-D, UV and IR spectra were recorded on JEOL JNM-ECP 600, JASCO J-810, JASCO P-1020 digital, Beckman DU® 640 and Nicolet NEXUS 470 spectrophotometers, respectively. ESI-MS, EI-MS and GC-MS were measured on Q-TOF ULTIMA GLOBAL GAA076 LC, VG Autospec-3000 and Agilent 6890/5973 spectrometers, respectively. Semipreparative HPLC and chiral separation was performed on a YMC-pack ODS-A column  and a CHIRALPAK IA column . TLC was performed on plates precoated with silica gel GF254 (10–40 μm). The column chromatography (CC) was performed over silica gel  and Sephadex LH-20 , respectively. The seawater for the cultural medium of Cladosporium sp. OUCMDZ-302 was collected from Yellow Sea near Qingdao.The NMR, ECD, [α]Cladosporium sp. OUCMDZ-302 was isolated from the surface sterilized stems of the mangrove plant E. agallocha grown in Wenchang, Hainan, China. Briefly, the stems were washed with tap water and sterile distilled water in sequence. The stems with clean surface were further sterilized in a sequence of 75% ethanol for 2 min, 0.1% of HgCl2 for 3 min, and sterile distilled water. The outer bark was removed, and the inner bark was cut into small pieces that were then placed on a potato dextrose agar (PDA) plate and cultured at 28°C for 3 days. A single colony was transferred to PDA media and was identified according to its morphological characteristics  and cultured at 28°C for 48 h on a rotary shaker at 120 rpm. The seed culture was transferred into three hundred and fifty 500 mL conical flasks (200 mL/flask) containing the same medium, and performed at 28°C for 7 days on rotary shakers at 160 rpm. The whole fermentation broth (70 L) was filtered through cheese cloth to separate the mycelia from filtrate. The filtrate was concentrated to about one-quarter of the original volume under reduced pressure and then extracted three times with equal volumes of ethyl acetate (EtOAc) and concentrated to dryness. The mycelia were extracted three times with acetone and concentrated to an aqueous solution. The aqueous solution was subsequently extracted three times with equal volumes of EtOAc and concentrated. Both EtOAc extractions were combined to give 45 g of the extract.The producing fungal strain 3-petroleum ether  and then MeOH–CHCl3 . Fr.1 (5.4 g) was separated on a silica gel column  eluted with CHCl3-petroleum ether  to give 12 (1g). Fr.3 (0.3 g) was further purified by semipreparative HPLC (60% MeOH/H2O) to give 10 . Fr.4 (4.3 g) was separated into two subfractions by column chromatography over silica gel (RP-18) eluting with gradient H2O-MeOH (50–100%). Fr.4-1 (1.4 g) was separated by Sephadex LH-20  to obtain three fractions . Fr.4-1-2 (140 mg) was purified by semipreparative HPLC (30% MeOH/H2O) to yield 5 , 13 , and 15 . Fr.4-1-3 (190 mg) was purified by semipreparative HPLC  to give 4  and 8 . Fr.4-2 (360 mg) was purified by semipreparative HPLC (50% MeOH/H2O) to give 6 , and 14 . Fr.5 (1.1 g) was separated into two subfractions by a silica gel column  eluted with MeOH–CHCl3 . Fr.5-1 (40 mg) was purified by semipreparative HPLC (25% MeOH/H2O) to give 3 , and Fr.5-2 (80 mg) was purified by semipreparative HPLC  to give compounds 11 . Fr.6 (2.8 g) was separated into two subfractions by a silica gel column  eluted with CHCl3-petroleum ether MeOH–CHCl3 . Fr.6-1 (110 mg) was purified by semipreparative HPLC (60% MeOH/H2O) to give 9 . Fr.6-2 (340 mg) was purified by semipreparative HPLC (50% MeOH/H2O) to give 7  and the mixture of 1 and 2 . The mixture of 1 and 2 were further purified by a chiral column  to yield compound 1  and 2 .The extract (45 g) was separated into eight fractions (Fr.1–Fr.8) on a silica gel column  using a step gradient elution with CHClD were performed in HyperChem 7.5 and Gaussian 03  from the proton-proton torsion angle , indicating seven degrees of unsaturation. An interpretation of the 1D (Table 11) -11 and D-ribose that was identified by GC-MS analysis of the reaction products with L-cysteine methyl ester and Me3SiCl - and (2S)-7-O-α-D- ribofuranosyl-5-hydroxy-2-methylchroman-4-one, respectively. This is the first time that to solidify the absolute configuration of compound 1, although it was reported last year , indicating five degrees of unsaturation. The NMR data  to H-4 (δ 4.48) through H-2 (δ 4.14) and H-3 (δ 3.45)  is consistent with the calculated one for -3 (−100) and opposite to the calculated one for -3 (+102). Thus, the structure of 3 was identified as -2-methylchroman-3,4,5-triol.The molecular formula of compound ta Table –S14 was ) Figure , and the) Figure . In orde4 showed the molecular formulae of C11H14O5 based on HRESIMS peaks at m/z 225.0767 [M–H]− , indicating five degrees of unsaturation. The 1D . The 1D NMR data  and the upfield shift for carbonyl carbon (−3.5 ppm), indicating that 4 is the methyl ester of 8. This was confirmed by analysis of 1H-1H COSY correlation  and 8 ([α]D +8.2) were opposite to the synthetic analog, R-3-(3-methoxyphenyloxy)butanoic acid ([α]D −31.2) -3- butanoate and (3S)-3-butanoic acid, respectively.Compound  Figures –S19 and  Figures  NMR specn Figure  and the 0 Figure . The spe5 was determined as C7H12O2 based on the HREIMS peak at m/z 128.0845 [M]+ , corresponding to two degrees of unsaturation. The IR spectrum showed hydroxy groups at 3442 cm−1 and double bonds at 3080 and 1646 cm−1. The 1D NMR spectra –CH (OH)–CH3 on the basis of 1H-1H COSY correlations from the methyl (δH 1.01) to the methylene (δH 5.03/5.17) through the two oxygenated methines  corresponded to E-Δ4 double bond. The large 3JH−2, H−3 value (6.0 Hz) and the downfield methyl carbon signal (δC−1 19.1) indicated an anti-conformation  matched with the measured one (6.0 Hz) while the calculated one of erythro-5 (3.8 Hz) was inconsistent, indicating threo- configuration. The direction of the specific rotation of 5 ([α]D −6.8) were similar to the structurally related t-butyl -6,7-dihydroxyocta-2,4-dienoate ([α]D −23) -6,7-dihydroxyocta-2,4-dienoate ([α]D +22.9) -hepta-4,6-diene-2,3-diol.The molecular formula of compound ra Table –S25 of 5s Figure . The lar6 was determined to be C11H18O2 based on the HREIMS peak at m/z 182.1305 [M]+ , indicating three degrees of unsaturation. The EIMS of 6 illustrated in Figure 2OH, -C6H9O, and -C5H9O moieties. The 1H –CH2-CH = CH–CH2-CH2OH, which was supported by HMBC correlations  and 3JH−8, H−9 (14.8 Hz) suggested that both Δ3 and Δ8 double bonds were E-configurations. The direction of specific rotation of 6 ([α]D +2.0) is similar to that of (S)-dodeca-3,5-diene-1,7-diol ([α]D +56) -undeca-3,8,10-triene-1,6-diol.The molecular formula of compound H Figure  and 13C C Figure  NMR specC Figure  and HMQCC Figure  experimeY Figure  correlats Figure . The lar9 and 10 were determined as (–)-trans-4-methoxy-2-methylchroman-5-ol -9 and opposite to ECD of -9 -4-methoxy-2-methyl chroman-5-ol. The similar sign of the specific rotations of 9 and 10 -2-methylchroman-4,5-diol. The absolute configurations of compounds 9 and 10 were determined for the first time in this study.The relative configurations of compounds 9 Figure . Thus, cR)-7-O-α-D-Ribofuranosyl-5-hydroxy-2-methylchroman-4-one (1): White amorphous powder; c 0.1, MeOH); UV (MeOH) λmax (log ε) 204 (3.62), 278 (3.55), 320 (2.77) nm; ECD (MeOH) λmax (Δε) 211 (+14.21), 284 (−3.51), 327 (+3.42); IR (KBr) νmax 3,416, 1,646, 1,573, 1,354, 1,295, 1,195, 1,155, 1,076, 1,029 cm−1; 1H and 13C NMR .(2an-4-one : White aS)-7-O-α-D-Ribofuranosyl-5-hydroxy-2-methylchroman-4-one (2): White amorphous powder; c 0.1, MeOH); UV (MeOH) λmax (log ε) 204 (3.62), 278 (3.55), 320 (2.77) nm; ECD (MeOH) λmax (Δε) 211 (−10.04), 284 (+9.40), 330 (−2.13); IR (KBr) νmax 3,416, 1,646, 1,573, 1,354, 1,295, 1,195, 1,155, 1,076, 1,029 cm−1; 1H and 13C NMR .-2-Methylchroman-3,4,5-triol (3): Colorless oil; c 0.1, MeOH); UV (MeOH) λmax (log ε) 200 (3.25), 270 (2.26) nm; IR (KBr) νmax 3,429, 2,356, 1,627, 1,4,01, 1,090 cm−1; 1H and 13C NMR .-3-butanoate (4): Colorless oil; c 0.1, MeOH); UV (MeOH) λmax (log ε) 200 (3.32), 270 (2.19) nm; IR (KBr) νmax 3,409, 2,356, 1,706, 1,606, 1,481, 1,202, 1,063, 1,010 cm−1; 1H and 13C NMR Methyl -Hepta-4,6-diene-2,3-diol (5): Colorless oil; c 0.1, MeOH); UV (MeOH) λmax (log ε) 200 (3.09), 217 (3.38) nm; IR (KBr) νmax 3442, 3080, 1646, 1540, 1023, 446 cm−1; 1H and 13C NMR (Table m/z (%): 129 (45), 256 (8), 111 (26), 97 (51), 83 (69), 82 (38); HREIMS m/z 128.0845 [M]+ .2S,3S,4E-HE,8E,6S)-Undeca-3,8,10-trien-1,6-diol (6): Colorless oil; c 0.1, MeOH); UV (MeOH) λmax (log ε) 200 (2.86), 218 (3.16) nm; IR (KBr) νmax 3,390, 2,927, 1,715, 1,421, 1,047, 973 cm−1; 1H and 13C NMR (Table m/z (%): 181 (8), 164 (9), 151 (14), 129 (17), 115 (16), 97 (31), 85 (28), 71 (53), 67(88), 53(10); HREIMS m/z 182.1305 [M]+ . Å, b = 10.6659(11) Å, c = 9.8560(8) Å, α = 90.00°, β = 95.3230(10)°, γ = 90.00°, V = 733.95(12) Å3, Z = 2, Dcalcd = 1.476 mg/m3, μ = 0.121 mm−1, F(000) = 344, crystal size 0.42 ×0.30 ×0.21 mm. A total of 3413 unique reflections (2θ<50°= were collected on a CCD area detector diffractometer with graphite monochromated Mo-Ka radiation (λ = 0.71073 Å). The structure was solved by direct methods (SHELXS-97) and expanded using Fourier techniques (SHELXL-97). The final cycle of full-matrix least squares refinement was based on 2053 unique reflections (2θ <50°) and 210 variable parameters and converged with unweighted and weighted agreement factors of R1 = 0.0421, Rw = 0.0981 and R = 0.0374 for I>2sigma(I) data. Crystallographic data (excluding structure factors) for structure 1 in this paper have been deposited in the Cambridge Crystallographic Data Centre as supplementary publication number CCDC 883328 [fax: +44 (0)-1223-336033 or e-mail: deposit@ccdc.cam.ac.uk].Compound 11 and 12. Compound 11 formed compounds 1 and 2 by glycosidation. (S)-12 underwent oxidation and reduction to yield compound 3. The reduction of (S)-12 produced compound 10 that was transformed to compound 9 followed by methylation. (S)-12 was subjected to Baeyer-Villiger oxidation followed by methanolysis and hydrolysis to yield compounds 4 and 8, respectively. Compounds 5 and 6 were formed from different lengths of acetyl-CoA units by condensation, reduction, dehydration, and decarboxylation. The condensation of acetyl-CoA units followed by cyclization and reduction formed compound 15 that was transformed to compound 14 after enolization and dehydration.These compounds were postulated to be biosynthesized by the polyketide pathway from acetyl coenzyme A Figure . The ace1–14 were tested for cytotoxic effects on the HL-60, BEL-7402, K562, A549, HeLa, and H1975 cell lines, DPPH scavenging activity, and antimicrobial activities against E. coli, E. aerogenes, P. aeruginosa, B. subtilis, and C. albicans. As the results, compound 6 was cytotoxic to H1975 cell line with an IC50 values of 10.0 μM, while compounds 4 and 8–10 showed DPPH radical scavenging activity with the IC50 values of 2.65, 0.24, 5.66, and 6.67 μM, respectively. None of the compounds exhibit antimicrobial activities.Compounds Cladosporium sp. OUCMDZ-302 with Excoecaria agallocha. The new compound 4 showed DPPH radical scavenging activity with an IC50 value of 2.65 μM.Five new polyketides were isolated and identified from the fermentation of the mangrove fungus All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


H-1,2,3-triazol-4-yl)ethylidene)hydrazine-1-carbodithioate and 2-ethylidene)hydrazine-1-carbothioamide afforded 2-ethylidene)hydrazono)-3-phenyl-5-substituted-2,3-dihydro-1,3,4-thiadiazoles and 5-(4-substituted)diazenyl)-2-ethylidene)hydrazinyl)-4-arylthiazoles, respectively. Analogously, the reactions of hydrazonoyl halides with 7--5-phenyl-2-thioxo-2,3-dihydropyridopyrimidin-4(1H)-one gave 3-(4-substituted)-8--6-phenyl-1-arylpyrido--triazolo-pyrimidin-5(1H)-ones in a good yield. The structures of the newly synthesized were elucidated via elemental analysis, spectral data and alternative synthesis routes whenever possible. Twelve of the newly synthesized compounds have been evaluated for their antitumor activity against human breast carcinoma (MCF-7) and human hepatocellular carcinoma (HepG2) cell lines. Their structure activity relationships (SAR) were also studied. The 1,3,4-thiadiazole derivative 9b (IC50 = 2.94 µM) has promising antitumor activity against the human hepatocellular carcinoma cell line and the thiazole derivative 12a has promising inhibitory activity against both the human hepatocellular carcinoma cell line and the breast carcinoma cell line .Reactions of hydrazonoyl halides and each of methyl 2-(1-(5-methyl-1-phenyl-1 Several1,3,4-Thiadiazole derivatives have attracted considerable interest owing to their wide spectra of biological activities such as antibacterial, antifungal, antituberculosis, antihepatitis B viral, antileishmanial, anti-inflammatory, analgesic, CNS depressant, anticancer, antioxidant, antidiabetic, molluscicidal, antihypertensive, diuretic, analgesic, antimicrobial, antitubercular, and anticonvulsant activities ,24,25,26Thiazoles can found in drug development for the treatment of allergies , hyperted]triazolopyrimidine derivatives containing 1,2,3-triazole moieties.The 1,2,4-triazolopyrimidines have also attracted growing interest due to their important pharmacological activities, such as antitumor potency, antimalarial, antimicrobial, anti-inflammatory, antifungal and macrophage activation ,40,41,42H-1,2,3-triazole (1)  to the C=S double bond of 3. The formation of 6 and 7 are similar to the reactions of hydrazonoyl chloride with 1-phenyl-1,4-dihydrotetrazole-5-thione pyrimidin-4(1H)-one (17) in a good yield. Structure 17 was elucidated by elemental analysis, spectral data and chemical transformation. Thus, when compound 17 was reacted with 4a in chloroform under reflux it afforded one isolable product, as evidenced by tlc, formulated as 3-acetyl-8--1,6-diphenylpyridotriazolopyrimidin-5(1H)-one  1,3-addition of the thiol tautomer 18 to the nitrilium imide 5 to give the thiohydrazonate ester 19 which undergoes nucleophilic cyclization to yield spiro compounds 20. The latter ring opened and cyclized to yield 22 by loss of hydrogen sulfide; and (2) 1,3-cycloaddition of nitrilium imide 5 to the C=S double bond of 17 to give 20 directly   was reacone 22a, . The mecdirectly . Attempt50 and IC50 values, as depicted in Our literature survey showed that many thiazole and 1,3,4-thiadiazole derivatives have antitumor activity with excellent IGin vitro growth inhibitory activity of the synthesized compounds was investigated in comparison with the well-known anticancer standard drug doxorubicin using a crystal violet colorimetric viability assay. Data generated were used to plot a dose response curve of which the concentration of test compounds required to kill 50% of the cell population (IC50) was determined. The cytotoxic activity was expressed as the mean IC50 of three independent experiments  have the following descending order: 12a > 9c > 9b > 9g > 12e > 9d > 12b > 9e > 9a > 22d > 9f > 22a.The -in vitro inhibitory activities of tested compounds against the human hepatocellular carcinoma (HepG2) cell line have the following descending order: 12a > 9b > 9g > 9d > 9c > 9e > 9a > 12e > 22d > 12b > 9f > 22a.The The eriments  and the -9b (IC50 = 2.94 µM) has promising antitumor activity against the human hepatocellular carcinoma cell line while the other 1,3,4-thiadiazole derivatives 9a, 9c–f have moderate activities (IC50 = 7.72‒43.43 µM).The 1,3,4-thiadiazole -12a has promising inhibitory activity against both the human hepatocellular carcinoma cell line and the breast carcinoma cell line  while the other thiazole derivatives 12b and 12e have moderate activity.Thiazole -22a,d have moderate activity.Pyridotriazolopyrimidinone derivatives -in vitro inhibitory activity of tested compounds against the human breast carcinoma cell line have the following descending order: CONHC6H5 > COOC2H5 > C6H5 > C6H5CO > C10H7CO > CH3CO > C4H3SCO group.For substituents at position 2 of the 1,3,4-thiadiazole ring, the -in vitro inhibitory activity of tested compounds against the human hepatocellular carcinoma cell line have the following descending order: COOC2H5 > C6H5 > C6H5CO > CONHC6H5 > C10H7CO > CH3CO > C4H3SCO group.For substituents at position 2 of the 1,3,4-thiadiazole ring, the Examination of the SAR leads to the following conclusions:1H-NMR) and run in deuterated dimethylsulfoxide (DMSO-d6). Chemical shifts were related to that of the solvent. 13C-NMR was recorded on a Bruker spectrometer at 75 MHz. Mass spectra were recorded on a Shimadzu GCMS-QP1000 EX mass spectrometer at 70 eV. Elemental analyses were measured by using an ElementarVario LIII CHNS analyzer. Antitumor activity of the productswas carried out at the Regional Center for Mycology and Biotechnology at Al-Azhar University, Cairo, Egypt. Hydrazonoyl halides 4 triazolo- pyrimidin-5(1H)-one (22a). Yellow solid, (82% yield), mp 262–264 °C; IR: ν = 1670, 1651 (2C=O), 1599 (C=N) cm−1; 1H-NMR: δ = 2.42 , 2.68 ,7.30–8.38 ; MS, m/z (%) 538 , 373 (18), 260 (23), 156 (21), 80 (100), 56 (23). Anal. Calcd. forC31H22N8O2 (538.56): C, 69.13; H, 4.12; N, 20.81. Found: C, 69.08; H, 4.02; N, 20.68%.3-Acetyl-8--6-phenyl-1-(p-tolyl)pyridotriazolo pyrimidin-5(1H)-one (22b). Yellow solid, (76% yield), mp 262–264 °C; IR: ν = 1721, 1670 (2C=O), 1601(C=N) cm−1; 1H-NMR: δ = 2.26 , 2.41 , 2.64 , 7.14–7.99 ; MS, m/z (%) 552 , 515 (23), 370 (25), 217 (28), 106 (79), 52 (100). Anal. Calcd. For C32H24N8O2 (552.59): C, 69.55; H, 4.38; N, 20.28. Found: C, 69.38; H, 4.19; N, 20.21%.3-Acetyl-1-(4-chlorophenyl)-8--6-phenylpyrido triazolopyrimidin-5(1H)-one (22c). Yellow solid, (79% yield), mp 278–280 °C; IR: ν = 1721, 1670 (2C=O), 1600 (C=N) cm−1; 1H-NMR: δ = 2.42 , 2.64 , 7.19–7.97 ; MS, m/z (%) 573 , 213 (25), 129 (32), 98 (100), 57 (94). Anal. Calcd. For C31H21ClN8O2 (573.00): C, 64.98; H, 3.69; N, 19.56. Found: C, 64.75; H, 3.61; N, 19.44%.Ethyl 8--5-oxo-1,6-diphenyl-1,5-dihydropyrido triazolopyrimidine-3-carboxylate (22d). Yellow solid, (73% yield), mp 246–248 °C; IR: ν = 1749, 1670 (2C=O), 1601 (C=N) cm−1; 1H-NMR: δ = 1.29 , 2.40 , 4.13 , 7.17–7.64  ppm; MS, m/z (%) 568 , 481 (19), 236 (17), 111 (32), 69 (10), 55 (100). Anal. Calcd. For C32H24N8O3 (568.58): C, 67.60; H, 4.25; N, 19.71. Found: C, 67.43; H, 4.20; N, 19.58%.Ethyl 8--5-oxo-6-phenyl-1-(p-tolyl)-1,5-dihydropyrido triazolopyrimidine-3-carboxylate (22e). Yellow solid, (71% yield), mp 225–227 °C; IR: ν = 1750, 1653 (2C=O), 1601 (C=N) cm−1; 1H-NMR: δ = 1.32 , 2.24 , 2.41 , 4.16 , 7.12–7.83 ; MS, m/z (%) 582 , 431 (20), 222 (33), 131 (26), 76 (100). Anal. Calcd. For C33H26N8O3 (582.61): C, 68.03; H, 4.50; N, 19.23. Found: C, 68.17; H, 4.37; N, 19.07%.Ethyl 1-(4-chlorophenyl)-8--5-oxo-6-phenyl-1,5-dihydro-pyridotriazolopyrimidine-3-carboxylate (22f). Yellow solid, (77% yield), mp 270–272 °C; IR: ν = 1750, 1652 (2C=O), 1601 (C=N) cm−1; 1H-NMR: δ = 1.34 , 2.43 , 4.21 , 7.18–7.89 ; MS, m/z (%) 603 , 504 (67), 314 (85), 279 (73), 176 (95), 66 (100). Anal. Calcd. For C32H23ClN8O3 (603.03): C, 63.74; H, 3.84; N, 18.58. Found: C, 63.70; H, 3.75; N, 18.47%.8--5-oxo-N,1,6-triphenyl-1,5-dihydropyrido-triazolopyrimidine-3-carboxamide (22g). Yellow solid, (75% yield), mp 270–272 °C; IR: ν = 3325 (NH), 1668, 1651 (2C=O), 1601 (C=N) cm−1; 1H-NMR: δ = 2.24 , 7.13–8.06 , 11.27 ; 13C-NMR: δ = 10.3 (CH3), 120.1, 121.2, 121.3, 121.4, 121.8, 121.9, 125.2, 125.5, 125.8, 126.8, 127.8, 128.2, 128.8, 129.2, 129.6, 129.7, 130.2, 134.1, 135.7, 136.6, 138.0,138.4, 146.6, 148.8, 153.3 (Ar-C), 165.4, 173.6 (C=O); MS, m/z (%) 538 , 373 (18), 260 (23), 156 (21), 80 (100), 56 (23). Anal. Calcd. For C36H25N9O2 (615.64): C, 70.23; H, 4.09; N, 20.48. Found: C, 70.28; H, 4.02; N, 20.27%.3-Benzoyl-8--1,6-diphenylpyridotriazolopyrimidin-5(1H)-one (22h). Yellow solid, (78% yield), mp 255–257 °C; IR: ν = 1669, 1652 (2C=O), 1600 (C=N) cm−1; 1H-NMR: δ = 2.28 , 7.11–8.06 ; MS, m/z (%) 600 , 504 (32), 300 (40), 148 (87), 95 (59), 67 (100). Anal. Calcd. For C36H24N8O2 (600.63): C, 71.99; H, 4.03; N, 18.66. Found: C, 71.69; H, 4.01; N, 18.54%.2 and were subcultured two to three times a week.Human breast carcinoma (MCF-7) and human hepatocellular carcinoma (HepG2) cell lines were obtained from the American Type Culture Collection . The cells were grown on RPMI-1640 medium supplemented with 10% inactivated fetal calf serum and 50 µg/mL gentamycin. The cells were maintained at 37 °C in a humidified atmosphere with 5% CO4 cells adhered at the bottom of the wells in a 96-well microtiter plate incubated for 24 h at 37 °C in a humidified incubator with 5% CO2. The mono layers were then washed with sterile phosphate buffered saline  and simultaneously the cells were treated with 100 µL from different dilutions of tested sample in fresh maintenance medium and incubated at 37 °C. A control of untreated cells was made in the absence of tested sample. Positive control containing doxroubcin drug was also tested as reference drug for comparison. Six wells were used for each concentration of the test sample. Every 24 h the observation under the inverted microscope was made. The number of the surviving cells was determined by staining the cells with crystal violet [50), the concentration required to cause toxic effects in 50% of intact cells, was estimated from the graphic plots.Potential cytotoxicity of the compounds was evaluated on tumor cells using the method of Gangadevi and Muthumary . The cell violet ,67 follo12a has promising antitumor activities  and most of the tested compounds showed moderate anti-cancer activities.Some newly synthesized compounds were evaluated for their anti-cancer activity against the human breast carcinoma (MCF-7) and human hepatocellular carcinoma (HepG2) cell lines. Also, their structure activity (SAR) was studied. The results revealed that the thiazole derivative