Synthesis and Antifungal Activity of (Z)-/(E)-Verbenone Oxime Ether Compounds
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摘要:
将α-蒎烯选择性氧化制备马鞭草烯酮，对羰基进行肟化和分离，再发生亲核取代反应，合成得到40个新型（Z）-/（E）-马鞭草烯酮肟醚（4a~4t，包括20对Z/E异构体），采用1H NMR、13C NMR、FT-IR、UV-vis和ESI-MS对目标化合物进行了结构表征，并测试其抑菌活性。研究结果表明：在质量浓度50 mg/L下，目标化合物对8种植物病原菌均显示出不同程度的抑菌活性，其中化合物（E）-4r（R=2，6-Cl）对苹果轮纹病菌的抑制率为77.8%，化合物（E）-4s（R=2，6-F）对水稻纹枯病菌的抑制率为72.7%，化合物（E）-4n（R=p-CN）对玉米小斑病菌的抑制率为70.8%，（Z）-/（E）-异构体对一些植物病原菌的抑制活性显示一定差异。建立了（E）-马鞭草烯酮肟醚化合物对水稻纹枯病菌抑制活性的CoMFA模型（r2=0.992，q2=0.507），进行3D-QSAR研究，结果表明建立的模型可用于设计具有潜在高活性的先导化合物。
Abstract:Verbenone was prepared by selective oxidation of α-pinene at first. Then, by oximation of the carbonyl group (Z)-and(E)-verbenone oxime compounds were synthesized and separaed, followed by the nucleophilic substitution reaction to afford forty novel (Z)-and (E)-verbenone oxime ether compounds 4a-4t, including twenty couples of Z/E isomers. The structures of the target compounds were characterized by 1H NMR, 13C NMR, FT-IR, UV-vis, and ESI-MS. The antifungal activities of the target compounds were evaluated. The results showed that, at the mass concentration of 50 mg/L, all the target products exhibited certain inhibition activities against the eight kinds of plant pathogens. Compound(E)-4r(R=2, 6-Cl) had inhibition rates of 77.8% against Physalospora piricola. Compound(E)-4s(R=2, 6-F) had inhibition rates of 72.7% against Rhizoctonia solani. Compound (E)-4n(R=p-CN) had inhibition rates of 70.8% against Helmithosporium maydis. Certain difference were found in inhibition activity of Z-E isomers against some plant pathogens. The CoMFA model(r2=0.992, q2=0.507) of (E)-verbenone oxime ether compounds for inhibiting Rhizoctonia solani was established for the study of 3D-QSAR, and the results could provide a basis for the design of potential lead compounds with higher activity.
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Keywords:
- α-pinene /
- verbenone /
- oxime ether /
- Z-E isomer /
- antifungal activity /
- 3D-QSAR
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马鞭草烯酮[1]是具有一定生物活性的双环单萜烯酮类化合物，有类似于樟脑、薄荷脑以及芹菜的香气，常被用作香料及医药中间体。马鞭草烯酮存在于天然产物中，如蓝桉和西班牙马鞭草精油，以及长期存放的被氧化的松节油中, 也可由松节油主成分α-蒎烯的烯丙位亚甲基氧化得到[2-3]。马鞭草烯酮自身具有驱虫[4-5]和抗惊厥[6]活性，在农药和医药领域有广泛应用，其衍生物可用作不对称合成[7]、氨基酸类药物合成的起始物[8-9]和医药中间体[10]等。肟醚类化合物具有抗菌[11-12]、抗肿瘤[13-14]、杀螨[15]、杀虫[16]和酶抑制[17]等生物活性。基于大量文献报道及课题组多年来在松香松节油基生物活性化合物的研究成果[18-20]，作者将肟醚活性基团引入到马鞭草烯酮骨架中，合成系列具有潜在生物活性的(Z)-/(E)-马鞭草烯酮肟醚化合物，以期为我国天然优势生物质资源松节油的深度开发和利用提供新的途径。
1. 实验
1.1 试剂与仪器
α-蒎烯(GC纯度98.0%)，广西梧州松脂股份有限公司；盐酸羟胺、乙醇钠以及系列苄氯(苄溴)均为市售分析纯试剂，上海阿拉丁试剂公司。
AVANCE III HD 600 MHz超导核磁共振波谱仪，瑞士BRUKER公司；NICOLET IS 50 FT-IR红外光谱仪，美国THERMO SCIENTIFIC公司；TSQ QUANTUM ACCESS MAX液相色谱-质谱联用仪，美国THERMO SCIENTIFIC公司；SHIMASZU UV-1800紫外光谱仪，日本SHIMASZU公司；WATERS 1525高效液相色谱仪，美国WATERS公司；AGILENT 6890气相色谱仪，美国AGILENT TECHNOLOGIES公司；海能MP420全自动熔点仪，济南海能仪器股份有限公司。
1.2 (Z)-/(E)-马鞭草烯酮肟醚的合成
1.2.1 合成路线
(Z)-/(E)-马鞭草烯酮肟醚(4a~4t)的合成路线如下所示：
1.2.2 中间产物的制备
参考文献[1-2]制备马鞭草烯酮(2)，其为淡黄色油状液体。参考前期工作[18-19]制备得到(Z)-/(E)-马鞭草烯酮肟(3)(白色固体)，并用NOESY鉴定(Z)-/(E)-异构体。
1.2.3 目标产物的合成
参考文献[11]方法，在50.0 mL两口瓶中分别加入(Z)-或(E)-马鞭草烯酮肟(0.5 g, 3.0 mmol)、乙醇钠(0.3 g, 3.6 mmol)，无水乙醇(15 mL)，室温搅拌1 h，缓慢滴加苄氯或苄溴(3.6 mmol)的乙醇溶液到反应混合物中。采用TLC监测反应过程，待反应完全后，旋转蒸发除去乙醇。再加入适量去离子水，用乙酸乙酯萃取反应液3次，将有机层合并，再旋转蒸发除去溶剂。经柱层析纯化，得到目标化合物(Z)-或(E)-马鞭草烯酮肟醚(4a~4t)，均为黄色液体。
1.3 结构表征
对目标产物进行结构表征。以CH2Cl2为溶剂，采用紫外可见分光光度计分析紫外吸收；采用KBr压片法测定FT-IR光谱；以CDCl3为溶剂，TMS为内标，用600 MHz核磁共振仪进行1H NMR和13C NMR分析；采用电喷雾电离源(ESI)在液相色谱-质谱联用仪上进行ESI-MS分析。
1.4 抑菌活性测试
参照文献[21]，在质量浓度50 mg/L下，采用离体法，测试目标产物的抑菌活性，计算相对抑菌率。活性分级指标为A级: ≥90%, B级: ≥70%~90%, C级：≥50%~70%, D级: < 50%。
1.5 3D-QSAR研究
参照文献[22-23]，采用SYBYL-X 2.1.1软件对(E)-马鞭草烯酮肟醚(4a~4t)进行建模。所有建模目标化合物均采用Tripos力场、Gasteiger-Huckel电荷、Conjugate Gradient法，在最大循环次数为10 000次、终止收敛能量为0.05 kJ/mol，分子力场计算以+1价sp3杂化的碳原子作为探针、步长为0.2 nm，其余参数均为缺省值条件下进行结构优化。对水稻纹枯病菌的抑制活性建立CoMFA模型，进行3D-QSAR研究。
2. 结果与讨论
2.1 (Z)-/(E)-马鞭草烯酮肟醚的表征
(E)-4f:(E)-马鞭草烯酮肟间-甲氧基苄基醚，淡黄色液体，产率75.4%。UV-vis(CH2Cl2) λmax: 267.62 nm; IR(KBr, cm-1) ν: 3054(＝CH), 2998, 2955, 2932, 2869, 2838(C—H), 1621, 1602, 1588, 1492, 1436(C＝N, Ar—C＝C, C＝C), 1266, 1041(C—O); 1H NMR(600 MHz, CDCl3) δ :7.25(dd, J=10.4, 5.3 Hz, 1H, H-16), 6.94(dd, J=13.0, 4.8 Hz, 2H, H-15, H-17), 6.82(dd, J=8.2, 2.3 Hz, 1H, H-13), 6.45(dd, J=3.1, 1.5 Hz, 1H, H-3), 5.07(s, 2H, H-11), 3.80(s, 3H, H-18), 2.69(td, J=6.0, 1.5 Hz, 1H, H-1), 2.64(dt, J=8.9, 5.5 Hz, 1H, H-5), 2.25~2.21(m, 1H, H-7), 1.90(d, J=1.6 Hz, 3H, H-10), 1.71(d, J=8.9 Hz, 1H, H-7), 1.41(s, 3H, H-9), 0.90(s, 3H, H-8); 13C NMR(151 MHz, CDCl3) δ：159.94(C-2), 159.63(C-4), 158.89(C-14), 140.07(C-12), 129.30(C-16), 120.01(C-17), 113.17(C-13), 113.07(C-15), 110.30(C-3), 75.22(C-11), 55.20(C-18), 49.36(C-1), 48.32(C-6), 48.09(C-5), 37.60(C-7), 26.13(C-8), 23.56(C-10), 21.83(C-9); ESI-MS m/z: 286.10[M+H]+。
(Z)-4n:(Z)-马鞭草烯酮肟对-氰基苄基醚，淡黄色液体，产率76.8%。UV-vis(CH2Cl2) λmax: 235.53 nm; IR(KBr, cm-1) ν: 3045(＝CH), 2977, 2955, 2931, 2870(C—H), 2229(C≡N), 1634, 1610, 1506, 1442(C＝N, Ar—C＝C, C＝C), 1242, 1017(C—O); 1H NMR(600 MHz, CDCl3) δ: 7.65~7.59(m, 2H, H-14, H-16), 7.41(dd, J=8.0, 0.5 Hz, 2H, H-13, H-17), 5.75(dd, J=3.2, 1.6 Hz, 1H, H-3), 5.12(s, 2H, H-11), 3.60(td, J=5.9, 1.6 Hz, 1H, H-1), 2.61(dt, J=9.0, 5.5 Hz, 1H, H-5), 2.29~2.19(m, 1H, H-7), 1.87(d, J=1.6 Hz, 3H, H-10), 1.66~1.60(m, 1H, H-7), 1.45(s, 3H, H-9), 0.88(s, 3H, H-8); 13C NMR(151 MHz, CDCl3) δ ：162.91(C-2), 155.13(C-4), 144.34(C-12), 132.10(C-14, C-16), 127.85(C-13, C-17), 118.94(C-18), 115.40(C-15), 111.13(C-3), 74.23(C-11), 49.08(C-1), 47.60(C-6), 42.42(C-5), 36.42(C-7), 26.12(C-8), 23.15(C-10), 22.26(C-9); ESI-MS m/z: 281.08[M+H]+。
(E)-4s:(E)-马鞭草烯酮肟2′, 6′-二氟苄基醚，淡黄色液体，产率76.9%。UV-vis(CH2Cl2) λmax: 257.31 nm; IR(KBr, cm-1) ν: 3076(＝CH), 2953, 2901, 2873(C—H), 1626, 1595, 1471(C＝N, Ar—C＝C, C＝C), 1237, 1026(C—O); 1H NMR(600 MHz, CDCl3) δ:7.26(ddd, J=8.4, 5.5, 2.0 Hz, 1H, H-15), 6.91~6.86(m, 2H, H-14, H-16), 6.33(dd, J=3.1, 1.5 Hz, 1H, H-3), 5.14(d, J=0.9 Hz, 2H, H-11), 2.68(td, J=6.0, 1.5 Hz, 1H, H-1), 2.64~2.61(m, 1H, H-5), 2.21~2.19(m, 1H, H-7), 1.86(d, J=1.6 Hz, 3H, H-10), 1.68(d, J=8.9 Hz, 1H, H-7), 1.40(s, 3H, H-9), 0.85(s, 3H, H-8); 13C NMR(151 MHz, CDCl3) δ：161.35(C-13, C-17), 160.21(C-2), 158.74(C-4), 129.99(C-15), 113.69(C-12), 111.25(C-14, C-17), 110.17(C-3), 62.81(C-11), 49.30(C-1), 48.29(C-6), 48.06(C-5), 37.54(C-7), 26.10(C-8), 23.48(C-10), 21.70(C-9); ESI-MS m/z: 292.08[M+H]+。
(Z)-4f:(Z)-马鞭草烯酮肟间-甲氧基苄基醚，淡黄色液体，产率76.5%。UV-vis(CH2Cl2) λmax: 257.31 nm; IR(KBr, cm-1) ν: 3045(＝CH), 2954, 2929, 2870, 2836(C—H), 1628, 1604, 1492, 1468, 1436(C＝N, Ar—C＝C, C＝C), 1264, 1045(C—O); 1H NMR(600 MHz, CDCl3) δ:7.24(dd, J=12.7, 4.9 Hz, 1H, H-16), 6.91(d, J=7.5 Hz, 1H, H-17), 6.89(s, 1H, H-13), 6.81(dd, J=8.2, 2.5 Hz, 1H, H-15), 5.79(dd, J=2.9, 1.4 Hz, 1H, H-3), 5.07(s, 2H, H-11), 3.79(s, 3H, H-18), 3.61(td, J=5.9, 1.5 Hz, 1H, H-1), 2.58(dt, J=8.9, 5.5 Hz, 1H, H-5), 2.25~2.19(m, 1H, H-7), 1.86(d, J=1.5 Hz, 3H, H-10), 1.61(d, J=8.9 Hz, 1H, H-7), 1.43(s, 3H, H-9), 0.88(s, 3H, H-8); 13C NMR(151 MHz, CDCl3) δ ：162.30(C-2), 159.61(C-4), 154.44(C-14), 140.05(C-12), 129.28(C-16), 119.97(C-17), 115.70(C-13), 113.12(C-15), 113.09(C-3), 75.29(C-11), 55.19(C-18), 49.10(C-1), 47.36(C-6), 42.40(C-5), 36.35(C-7), 26.10(C-8), 23.14(C-10), 22.25(C-9); ESI-MS m/z: 285.96[M+H]+。
(Z)-4r:(Z)-马鞭草烯酮肟2′, 6′-二氯苄基醚，淡黄色液体，产率76.8%。UV-vis(CH2Cl2) λmax: 256.16 nm; IR(KBr, cm-1) ν: 3045(＝CH), 2980, 2955, 2927, 2868(C—H), 1634, 1584, 1468, 1438(C＝N, Ar—C＝C, C＝C), 1244, 1034(C—O); 1H NMR(600 MHz, CDCl3) δ:7.32~7.28(m, 2H, H-14, H-16), 7.17(dd, J=8.4, 7.7 Hz, 1H, H-15), 5.78(dd, J=3.1, 1.5 Hz, 1H, H-3), 5.32(dd, J=29.0, 10.7 Hz, 2H, H-11), 3.53(td, J=5.9, 1.6 Hz, 1H, H-1), 2.53(dt, J=8.9, 5.5 Hz, 1H, H-5), 2.24~2.13(m, 1H, H-7), 1.85(d, J=1.6 Hz, 3H, H-10), 1.57(d, J=8.9 Hz, 1H, H-7), 1.38(s, 3H, H-9), 0.80(s, 3H, H-8); 13C NMR(151 MHz, CDCl3) δ：163.04(C-2), 154.73 (C-4), 137.13(C-12), 133.02(C-13, C-17), 129.78(C-15), 128.24(C-14, C-16), 115.64(C-3), 69.83(C-11), 49.05(C-1), 47.43(C-6), 42.20(C-5), 36.25(C-7), 26.04(C-8), 23.12(C-10), 22.06(C-9); ESI-MS m/z: 324.00[M+H]+。
在IR谱图中，C＝N、Ar—CC和C＝C伸缩振动的吸收峰出现在1650~1400 cm-1之间；C—O伸缩振动吸收峰出现在1250和1070 cm-1左右。在1H NMR谱图中，目标化合物(Z)-4a~(Z)-4t和(E)-4a~(E)-4t的马鞭草烯酮骨架中＝C—H的δ分别在5.7和6.4附近，马鞭草烯酮骨架上的饱和氢在δ3.6~0.5和δ3.0~0.5；在13C NMR谱图中，目标产物(Z)-4a~(Z)-4t和(E)-4a~(Z)-4t的C＝N双键化学位移分别在δ163和δ159左右，C＝C双键分别在δ162~113和δ159~110。质谱数据与目标化合物相对分子质量相符合。
本研究合成了一系列(Z)-/(E)-马鞭草烯酮肟醚(4a~4t)，选择化合物4f、4n、4r和4s进行表征。(其余化合物的表征数据见辅助材料)
(E)-4n:(E)-马鞭草烯酮肟对-氰基苄基醚，淡黄色液体，产率75.4%。UV-vis(CH2Cl2) λmax: 267.33 nm; IR(KBr, cm-1) ν: 3069(＝CH), 2953, 2931, 2868(C—H), 2230(C≡N), 1626, 1468, 1438(C＝N, Ar—C＝C, C＝C), 1244, 1017(C—O); 1H NMR(600 MHz, CDCl3) δ:7.64~7.60(m, 2H, H-14, H-16), 7.45(d, J=8.4 Hz, 2H, H-13, H-17), 6.43(dd, J=2.9, 1.4 Hz, 1H, H-3), 5.13(s, 2H, H-11), 2.68~2.62(m, 2H, H-1, H-5), 2.26(td, J=5.9, 1.3 Hz, 1H, H-7), 1.93(d, J=1.6 Hz, 3H, H-10), 1.78~1.69(m, 1H, H-7), 1.41(s, 3H, H-9), 0.89(s, 3H, H-8); 13C NMR(151 MHz, CDCl3) δ：160.65(C-2), 159.86(C-4), 144.42(C-12), 132.12(C-14, C-16), 127.83(C-13, C-17), 118.98(C-18), 111.08(C-15), 109.97(C-3), 74.11(C-11), 49.38(C-1), 48.33(C-6), 48.24(C-5), 37.69(C-7), 26.08(C-8), 23.65(C-10), 21.77(C-9); ESI-MS m/z: 281.11[M+H]+。
(E)-4r:(E)-马鞭草烯酮肟2′, 6′-二氯苄基醚，淡黄色液体，产率76.2%。UV-vis(CH2Cl2) λmax: 266.48 nm; IR(KBr, cm-1) ν : 3076(＝CH), 2955, 2929, 2870(C—H), 1625, 1594, 1471, 1437(C＝N, Ar—C＝C, C＝C), 1236, 1029(C—O); 1H NMR(600 MHz, CDCl3) δ 7.31(d, J=8.0 Hz, 2H, H-14, H-16), 7.20~7.13(m, 1H, H-15), 6.35(dd, J=2.9, 1.4 Hz, 1H, H-3), 5.37~5.30(m, 2H, H-11), 2.70(td, J=6.1, 1.5 Hz, 1H, H-1), 2.63(dt, J=8.9, 5.5 Hz, 1H, H-5), 2.27~2.18(m, 1H, H-7), 1.86(d, J=1.6 Hz, 3H, H-10), 1.69(t, J=10.8 Hz, 1H, H-7), 1.40(s, 3H, H-9), 0.87(s, 3H, H-8); 13C NMR(151 MHz, CDCl3) δ：160.55(C-2), 158.70(C-4), 137.20(C-12), 133.03(C-13, C-17), 129.80(C-15), 128.27(C-14, C-16), 110.26(C-3), 69.88(C-11), 49.32(C-1), 48.29(C-6), 48.15(C-5), 37.53(C-7), 26.13(C-8), 23.52(C-10), 21.71(C-9); ESI-MS m/z:324.02[M+H]+。
(Z)-4s:(Z)-马鞭草烯酮肟2′, 6′-二氟苄基醚，淡黄色液体，产率75.8%。UV-vis(CH2Cl2) λmax: 259.60 nm; IR(KBr, cm-1) ν: 3069(＝CH), 2952, 2929, 2871(C—H), 1628, 1595, 1471(C＝N, Ar—C＝C, C＝C), 1237, 1026(C—O); 1H NMR(600 MHz, CDCl3) δ:7.25(ddd, J=8.4, 7.2, 4.2 Hz, 1H, H-15), 7.01~6.73(m, 2H, H-14, H-16), 5.77(dd, J=3.1, 1.6 Hz, 1H, H-3), 5.14(s, 2H, H-11), 3.50(td, J=5.9, 1.6 Hz, 1H, H-1), 2.53(dt, J=9.0, 5.5 Hz, 1H, H-5), 2.20~2.15(m, 1H, H-7), 1.84(d, J=1.6 Hz, 3H, H-10), 1.56(d, J=9.0 Hz, 1H, H-7), 1.39(s, 3H, H-9), 0.80(s, 3H, H-8); 13C NMR(151 MHz, CDCl3) δ：162.99(C-13，C-17), 162.63(C-2), 154.56(C-4), 129.98(C-15), 115.67(C-3), 113.75(C-12), 111.18(C-14，C-17), 62.87(C-11), 49.03(C-1), 47.36(C-6), 42.19(C-5), 36.27(C-7), 26.02(C-8), 23.09(C-10), 22.04(C-9); ESI-MS m/z: 292.09[M+H]+。
2.2 抑菌活性测试结果
在质量浓度50 mg/L下，目标化合物(Z)-/(E)-4a~(Z)-/(E)-4t对所测8种植物病原菌均有一定的抑制活性，结果见表 1(文中仅列出4f、4n、4r和4s的抑菌活性，其余见辅助材料)。其中，(E)-4r(R=2, 6-Cl)对苹果轮纹病菌的抑制率为77.8%，与阳性对照百菌清相当；(E)-4s(R=2, 6-F)对水稻纹枯病菌的抑制率为72.7%；(E)-4n(R=p-CN)对玉米小斑病菌的抑制率为70.8%。此外，(E)-、(Z)-异构体对一些植物病原菌的抑制作用有一定的差异。例如，(E)-4f(R=m-OCH3)对苹果轮纹病菌的抑制率是(Z)-4f(R=m-OCH3)的5.5倍，抑制率分别为61.1%和11.1%。总体上，中间体马鞭草烯酮肟经过醚化作用后抑菌活性有所提高。
目标产物对不同菌种的抑菌率(50 mg/L)
Antibacterial rate of target products against different strains(50 mg/L)
% 化合物
compounds黄瓜枯萎病菌
Fusarium oxysporum花生褐斑病菌
Cercospora arachidicola苹果轮纹病菌
Physalos-pora piricola番茄早疫病菌
Alternaria solani小麦赤霉病菌
Gibberella zeae水稻纹枯病菌
Rhizoctonia solani玉米小斑病菌
Helmithos- porium maydis西瓜炭疽病菌
Colletot-richum lagenarium(Z)-4f 22.7 35.7 11.1 36.4 37.5 59.1 30.4 34.6 (E)-4f 18.2 14.3 61.1 36.4 25.0 57.6 26.1 19.2 (Z)-4n 23.8 35.7 66.7 63.2 45.0 57.5 29.2 29.4 (E)-4n 28.6 50.0 33.3 63.2 40.0 61.3 70.8 41.2 (Z)-4r 9.5 42.9 44.4 36.8 25.0 45.0 29.2 11.8 (E)-4r 18.2 28.6 77.8 9.1 41.7 68.2 39.1 42.3 (Z)-4s 19.0 50.0 50.0 52.6 45.0 55.0 41.7 35.3 (E)-4s 31.3 42.9 40.0 40.0 46.2 72.7 44.4 11.1 百菌清chlorothalonil 100.0 73.3 75.0 73.9 73.1 96.1 90.4 91.3 2.3 3D-QSAR研究
2.3.1 CoMFA模型
以图 1中*标记的原子作为公共叠合点，对优化后的目标化合物进行叠合和计算，叠合结果如图 2所示，计算CoMFA值见表 2。活性因子(ED)通过ED=lg{I/[(100-I)Mw] }计算得到(其中I为化合物对水稻纹枯病菌的抑制率，Mw为相对分子质量)。以CoMFA值作为自变量，活性因子ED作为因变量建立CoMFA模型。
化合物(E)-4a~(E)-4t的实验活性因子(ED)和预测活性因子(ED′)
The experimental factor(ED)and predicted factor (ED′)of compounds(E)-4a -(E)-4t
化合物compounds CoMFA值CoMFA value ED ED′ 残差residue (E)-4a(R=H) 120 -2.77 -2.72 -0.05 (E)-4b(R= o-CH3) 130 -2.38 -2.39 0.01 (E)-4c(R= m-CH3) 128 -2.96 -2.96 0.00 (E)-4d(R=p-CH3) 132 -2.19 -2.20 0.01 (E)-4e(R=o-OCH3) 132 -2.53 -2.52 -0.01 (E)-4f(R=m-OCH3) 132 -2.32 -2.32 0.00 (E)-4g(R=o-Cl) 128 -2.54 -2.50 -0.04 (E)-4h(R=m-Cl) 122 -2.44 -2.46 0.02 (E)-4i(R=p-Cl) 126 -2.49 -2.51 0.02 (E)-4j(R=o-NO2) 132 -3.25 -3.27 0.02 (E)-4k(R=m-NO2) 126 -2.43 -2.43 0.00 (E)-4l(R=p-NO2) 132 -2.41 -2.42 0.01 (E)-4m(R=m-F) 120 -2.25 -2.31 0.06 (E)-4n(R=p-CN) 126 -2.25 -2.25 0.00 (E)-4o(R=p-SCH3) 140 -2.35 -2.35 0.00 (E)-4p(R=p-vinyl) 132 -2.32 -2.32 0.00 (E)-4q(R=2, 6-CH3) 138 -2.53 -2.55 0.02 (E)-4r(R=2, 6-Cl) 132 -2.18 -2.17 -0.01 (E)-4s(R=2, 6-F) 122 -2.04 -2.04 0.00 (E)-4t(R=3-Cl-4-F) 124 -2.40 -2.36 -0.04 2.3.2 CoMFA的主成分分析
采用的留一法(Leave-one-out)交叉验证和非交叉验证对模型进行验证，相关验证参数如下：交叉验证相关系数(q2)0.507，非交叉验证相关系数(r2)0.992，估计标准误差(S)0.033，Fisher验证值(F)170，表明该模型具有可靠性和预测能力。主成分(PLS)分析显示，立体场和静电场的贡献分别为55.4%和44.6%，表明对活性的贡献主要是立体场。利用建立的CoMFA模型对建模的分子进行预测，其预测ED′值及残差值(ED值与ED′值之差)列于表 2。
以实验ED值为横坐标、预测ED′值为纵坐标作图，结果见图 3。图中实验值和预测值的数据都集中在45°对角线附近，进一步说明该模型的可靠性且具有较好的预测能力。
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图 3化合物(E)-4a~(E)-4t实测值与预测值的线性回归图
Figure 3.Linear regression of the experimental and predicted ED values of Compounds(E)-4a-(E)-4t
2.3.3 CoMFA等势图分析
![]()
图 4化合物(E)-4s的CoMFA模型的立体场(a)和静电场(b)分布图
Figure 4.3D contour plot of the CoMFA steric field(a) and electrostatic field(b) for compound(E)-4s
图 4所示即为(E)-4s模板分子的CoMFA三维等势图，其中(a)和(b)分别表示立体场三维等势图和静电场三维等势图。在图 4(a)中，绿色表示引入大体积基团会增强活性，黄色区域定义相反。在图(a)中，红色区域表示引入负电荷基团会增强活性，而蓝色区域定义相反。
在立体场图中，在苯环邻位和对位有大体积取代基有利于活性增加，例如化合物(E)-4e (R=o-OCH3)比(E)-4g(R=o-Cl)活性好；化合物(E)-4l(R=p-NO2)比(E)-4i(R=p-Cl)活性好。静电场图中，在苯环邻位上有吸电子取代基有利于活性增加，例如化合物(E)-4s(R=2, 6-F)>(E)-4r(R=2, 6-Cl)>(E)-4q(R=2, 6-CH3)。在苯环间位上有吸电子取代基也有利于活性增加，例如化合物(E)-4k(R=m-NO2)>(E)-4h(R=m-Cl)>(E)-4c(R=m-CH3)。在苯环对位上有推电子基团可增强其活性，例如，化合物(E)-4d(R=p-CH3)比(E)-4i(R=p-Cl)活性好。由此可知，建立的CoMFA模型可用于设计具有潜在高活性的先导化合物。
3. 结论
将α-蒎烯的烯丙位亚甲基进行选择性氧化制备马鞭草烯酮，再与盐酸羟胺发生肟化反应并分离，得到(Z)-/(E)-马鞭草烯酮肟，继而与系列苄氯或苄溴发生亲核取代反应，合成得到40个新型(Z)-/(E)-马鞭草烯酮肟醚化合物4a~4t(20对Z/E异构体)。利用1H NMR、13C NMR、FT-IR、UV-Vis和ESI-MS等多种波谱技术对目标产物做了结构表征。
抑菌活性测试表明，在质量浓度50 mg/L下，目标产物(Z)-/(E)-4a~(Z)-/(E)-4t对所测8种植物病原菌均有一定的抑制活性。其中，(E)-4r(R=2, 6-Cl)对苹果轮纹病菌的抑制率为77.8%；(E)-4s(R=2, 6-F)对水稻纹枯病菌的抑制率为72.7%；(E)-4n(R=p-CN)对玉米小斑病菌的抑制率为70.8%。此外，(Z)-、(E)-异构体对一些植物病原菌的抑制作用有一定的差异。
辅助材料(Supporting Information) 化合物4a~4t(除4f、4n，4r和4s)的表征及抑菌活性。这些材料可免费从本刊网站(http://www.cifp.ac.cn)下载。
建立了马鞭草烯酮肟醚化合物对抑制水稻纹枯病菌活性的CoMFA模型(r2=0.992, q2=0.507)，进行3D-QSAR研究，其结果可为设计具有潜在高活性的先导化合物提供依据。
致谢: -
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图 3
化合物(E)-4a~(E)-4t实测值与预测值的线性回归图
Figure 3.
Linear regression of the experimental and predicted ED values of Compounds(E)-4a-(E)-4t
![]()
图 4
化合物(E)-4s的CoMFA模型的立体场(a)和静电场(b)分布图
Figure 4.
3D contour plot of the CoMFA steric field(a) and electrostatic field(b) for compound(E)-4s
表 1
目标产物对不同菌种的抑菌率(50 mg/L)
Table 1
Antibacterial rate of target products against different strains(50 mg/L)
% 化合物
compounds黄瓜枯萎病菌
Fusarium oxysporum花生褐斑病菌
Cercospora arachidicola苹果轮纹病菌
Physalos-pora piricola番茄早疫病菌
Alternaria solani小麦赤霉病菌
Gibberella zeae水稻纹枯病菌
Rhizoctonia solani玉米小斑病菌
Helmithos- porium maydis西瓜炭疽病菌
Colletot-richum lagenarium(Z)-4f 22.7 35.7 11.1 36.4 37.5 59.1 30.4 34.6 (E)-4f 18.2 14.3 61.1 36.4 25.0 57.6 26.1 19.2 (Z)-4n 23.8 35.7 66.7 63.2 45.0 57.5 29.2 29.4 (E)-4n 28.6 50.0 33.3 63.2 40.0 61.3 70.8 41.2 (Z)-4r 9.5 42.9 44.4 36.8 25.0 45.0 29.2 11.8 (E)-4r 18.2 28.6 77.8 9.1 41.7 68.2 39.1 42.3 (Z)-4s 19.0 50.0 50.0 52.6 45.0 55.0 41.7 35.3 (E)-4s 31.3 42.9 40.0 40.0 46.2 72.7 44.4 11.1 百菌清chlorothalonil 100.0 73.3 75.0 73.9 73.1 96.1 90.4 91.3 
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表 2
化合物(E)-4a~(E)-4t的实验活性因子(ED)和预测活性因子(ED′)
Table 2
The experimental factor(ED)and predicted factor (ED′)of compounds(E)-4a -(E)-4t
化合物compounds CoMFA值CoMFA value ED ED′ 残差residue (E)-4a(R=H) 120 -2.77 -2.72 -0.05 (E)-4b(R= o-CH3) 130 -2.38 -2.39 0.01 (E)-4c(R= m-CH3) 128 -2.96 -2.96 0.00 (E)-4d(R=p-CH3) 132 -2.19 -2.20 0.01 (E)-4e(R=o-OCH3) 132 -2.53 -2.52 -0.01 (E)-4f(R=m-OCH3) 132 -2.32 -2.32 0.00 (E)-4g(R=o-Cl) 128 -2.54 -2.50 -0.04 (E)-4h(R=m-Cl) 122 -2.44 -2.46 0.02 (E)-4i(R=p-Cl) 126 -2.49 -2.51 0.02 (E)-4j(R=o-NO2) 132 -3.25 -3.27 0.02 (E)-4k(R=m-NO2) 126 -2.43 -2.43 0.00 (E)-4l(R=p-NO2) 132 -2.41 -2.42 0.01 (E)-4m(R=m-F) 120 -2.25 -2.31 0.06 (E)-4n(R=p-CN) 126 -2.25 -2.25 0.00 (E)-4o(R=p-SCH3) 140 -2.35 -2.35 0.00 (E)-4p(R=p-vinyl) 132 -2.32 -2.32 0.00 (E)-4q(R=2, 6-CH3) 138 -2.53 -2.55 0.02 (E)-4r(R=2, 6-Cl) 132 -2.18 -2.17 -0.01 (E)-4s(R=2, 6-F) 122 -2.04 -2.04 0.00 (E)-4t(R=3-Cl-4-F) 124 -2.40 -2.36 -0.04
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