董晶晶, 邵莉莉, 李金玉, 劉翰飛, 婁華勇, 潘衛(wèi)東,①

(1. 貴州大學(xué)藥學(xué)院, 貴州 貴陽 550025; 2. 貴州省天然產(chǎn)物研究中心, 貴州 貴陽 550014)

川楝(MeliaazedarachLinn.)為楝科(Meliaceae)楝屬(MeliaLinn．)植物,主要分布于四川、貴州和云南等地,具有行氣止痛和驅(qū)蟲等功效[1]。川楝的干燥根皮及樹皮是傳統(tǒng)中藥苦楝皮的重要來源,但有關(guān)川楝樹皮化學(xué)成分分離純化的研究尚不全面,川楝樹皮中化學(xué)成分結(jié)構(gòu)類型及其物質(zhì)活性有待進(jìn)一步探索。鑒于此,本文對川楝樹皮乙酸乙酯提取物開展了分離純化研究,以期為川楝樹皮的進(jìn)一步開發(fā)和利用提供基礎(chǔ)數(shù)據(jù)。

1 材料和方法

1.1 材料

于2019年9月,在貴州省興義市南盤江鎮(zhèn)采集株齡5 a川楝的樹皮,曬干后保存、備用。標(biāo)本由貴州黔西南喀斯特區(qū)域發(fā)展研究院的鄧超義教授鑒定。

1.2 方法

1.2.1 化合物分離 將曬干的川楝樹皮樣品(23 kg)粉碎成粗粉,用體積分?jǐn)?shù)95%乙醇加熱回流提取2次,每次2 h,減壓濃縮得到浸膏,浸膏用乙酸乙酯(分析純)萃取,得到乙酸乙酯部位浸膏(800 g)。對乙酸乙酯部位浸膏用聚酰胺拌樣,用體積分?jǐn)?shù)40%～100%甲醇(分析純)溶液梯度洗脫,分離得到6個流分(Fr.A～F),然后依次通過C18反相柱層析、硅膠柱層析、凝膠柱層析以及高效液相色譜儀純化后得到化合物1(500.0 mg)、化合物2(2.6 mg)、化合物3(31.8 mg)、化合物4(119.8 mg)、化合物5(9.2 mg)、化合物6(8.9 mg)、化合物7(11.1 mg)、化合物8(3.7 mg)、化合物9(2.4 mg)和化合物10(4.0 mg)。

1.2.2 高效液相條件 Agilent色譜柱(250 mm×9.4 mm,5 μm),柱溫28 ℃,流動相為體積分?jǐn)?shù)85%～45%的甲醇-水溶液,流速2 mL·min-1,檢測波長為210、230、254和280 nm。

2 結(jié)果和分析

化合物1:無色晶體,熔點(diǎn)163 ℃～164 ℃,分子式C30H44O3,ESI-MSm/z:475[M+Na]+。1H-NMR(600 MHz,CDCl3)δH:5.30(1H,d,J=3.0 Hz,H-7),5.06(1H,m,H-24),4.12(1H,m,H-16),2.72(2H,m,H-2),2.46(1H,m,H-9),2.38(1H,ddd,J=12.5,8.1,4.4 Hz,H-20),2.25(2H,m,H-15),2.21(2H,m,H-6),2.11(2H,d,J=3.9 Hz,H-11),1.99(2H,m,H-23),1.92(2H,dd,J=11.3,6.3,3.7 Hz,H-12),1.75(2H,m,H-22),1.69(1H,m,H-5),1.65(3H,s,H-27),1.57(3H,s,H-26),1.42(2H,m,H-1),1.21(3H,s,H-30),1.08(3H,s,H-29),1.01(3H,s,H-28),0.99(3H,s,H-18),0.92(3H,s,H-19)。13C-NMR(150 MHz,CDCl3)δC:216.1(C-3),180.5(C-21),143.3(C-8),132.5(C-25),123.3(C-24),118.3(C-7),82.3(C-16),58.0(C-17),55.0(C-14),52.4(C-5),47.7(C-9),47.7(C-4),45.3(C-20),39.4(C-13),38.1(C-1),35.5(C-15),35.3(C-10),34.6(C-2),32.1(C-30),29.5(C-22),29.1(C-12),25.9(C-6),25.6(C-27),24.3(C-11),24.2(C-28),21.4(C-29),21.3(C-19),17.8(C-26),16.7(C-23),12.3(C-18)。與文獻(xiàn)[2]比對,鑒定化合物1為苦內(nèi)酯(kulactone)。

化合物2:無色晶體,熔點(diǎn)203 ℃～204 ℃,分子式C30H44O4,ESI-MSm/z:491[M+Na]+。1H-NMR(600 MHz,CDCl3)δH:5.68(1H,d,J=2.8 Hz,H-7),5.09(1H,m,H-24),4.14(1H,m,H-16),3.35(1H,s,H-3),3.03(1H,m,H-9),2.57(1H,s,H-5),2.43(2H,m,H-20),2.32,1.79(2H,m,H-15),2.13(1H,m,H-17),2.11,2.03(2H,m,H-23),1.96,1.94(2H,m,H-11),1.92,1.86(2H,m,H-12),1.87,1.78(2H,m,H-2),1.82,1.34(2H,m,H-1),1.68(3H,s,H-27),1.61(3H,s,H-26),1.48(3H,m,H-22),1.26(3H,s,H-30),1.25(3H,s,H-28),1.13(3H,s,H-29),0.97(3H,s,H-18),0.86(3H,s,H-19)。13C-NMR(150 MHz,CDCl3)δC:200.8(C-6),180.1(C-21),167.3(C-8),132.8(C-25),124.6(C-7),123.2(C-24),81.6(C-16),76.5(C-3),60.7(C-5),57.6(C-17),55.9(C-14),49.7(C-9),45.2(C-20),44.1(C-10),39.2(C-4),36.7(C-13),34.8(C-15),30.9(C-1),29.6(C-30),29.1(C-22),29.0(C-12),27.9(C-28),26.0(C-23),25.6(C-27),24.4(C-2),21.4(C-29),21.3(C-18),17.9(C-26),16.4(C-11),14.0(C-19)。與文獻(xiàn)[3]比對,鑒定化合物2為3α-hydroxytirucalla-7,24(25)-dien-6-oxo-21,16-olide。

化合物3:白色粉末,分子式C30H44O4,ESI-MSm/z:491[M+Na]+。1H-NMR(600 MHz,CDCl3)δH:5.70(1H,d,J=2.8 Hz,H-7), 5.09(1H,t,J=7.1,1.5 Hz,H-24),4.15(1H,ddd,J=11.3,10.1,7.7 Hz,H-16),3.45(1H,s,H-5),3.20(1H,dd,J=11.7,3.6 Hz,H-3),2.90(1H,m,H-9),2.44(1H,ddd,J=12.5,7.9,4.6 Hz,H-20),2.32,1.76(2H,dd,J=13.8,10.1 Hz,H-15),2.12(1H,m,H-17),2.03(2H,m,H-23),1.94,1.48(2H,m,H-22),1.92,1.64(2H,m,H-11),1.82(2H,m,H-12),1.68(3H,s,H-26),1.66,1.39(2H,m,H-1),1.66,1.57(2H,m,H-2),1.61(3H,s,H-27),1.30(3H,s,H-28),1.26(3H,s,H-30),1.10(3H,s,H-29),0.98(3H,s,H-18),0.85(3H,s,H-19)。13C-NMR(150 MHz,CDCl3)δC:199.2(C-6),180.0(C-21),167.2(C-8),132.7(C-25),124.6(C-7),123.1(C-24),81.5(C-16),78.7(C-3),65.6(C-5),57.5(C-17),55.8(C-14),49.8(C-9),45.1(C-20),44.3(C-10),39.2(C-13),37.9(C-4),36.6(C-1),34.7(C-15),29.5(C-30),29.0(C-22),28.9(C-12),28.2(C-28),26.3(C-2),25.9(C-23),25.6(C-26),21.3(C-18),17.8(C-27),16.4(C-11),14.7(C-29),13.8(C-19)。與文獻(xiàn)[4]比對,鑒定化合物3為meliasenin B。

化合物4:白色粉末,分子式C30H42O4,ESI-MSm/z:489[M+Na]+。1H-NMR(600 MHz,CDCl3)δH:5.73(1H,t,J=2.4 Hz,H-7),5.05(1H,t,J=5.4 Hz,H-24),4.13(1H,m,H-16),2.94(1H,ddd,J=11.3,7.8,2.8 Hz,H-9),2.71,2.28(2H,m,H-2),2.70(1H,m,H-20),2.41(1H,s,H-5),2.32,1.89(2H,m,H-15),2.13(1H,d,J=12.0 Hz,H-17),2.06,2.00(2H,m,H-23),1.95,1.70(2H,m,H-1),1.94,1.45(2H,m,H-22),1.91,1.75(2H,m,H-11),1.82,1.66(2H,m,H-12),1.64(3H,s,H-27),1.57(3H,s,H-26),1.31(3H,s,H-29),1.29(3H,s,H-28),1.29(3H,s,H-30),1.07(3H,s,H-19),0.95(3H,s,H-18)。13C-NMR(150 MHz,CDCl3)δC:214.0(C-3),197.6(C-6),179.7(C-21),167.5(C-8),132.6(C-25),124.3(C-7),123.0(C-24),81.2(C-16),65.3(C-5),57.4(C-17),55.8(C-14),48.9(C-9),46.8(C-4),44.9(C-20),43.6(C-10),39.1(C-13),37.0(C-1),34.6(C-15),33.7(C-2),29.4(C-30),28.9(C-22),28.7(C-12),25.8(C-23),25.5(C-27),24.9(C-28),21.4(C-29),21.1(C-18),17.7(C-26),16.4(C-11),13.3(C-19)。與文獻(xiàn)[5]比對,鑒定化合物4為sendanolactone。

化合物5:無色油狀,分子式C30H42O4,ESI-MSm/z:491[M+Na]+。1H-NMR(600 MHz,CDCl3)δH:5.38(1H,d,J=3.0 Hz,H-7), 5.10(1H,t,J=6.6 Hz,H-24),4.18(2H,ddd,J=11.6,10.2,7.3 Hz,H-16),4.02(2H,dd,J=9.4,5.4 Hz,H-12),2.77(2H,td,J=14.6,5.6 Hz,H-2),2.52(1H,dd,J=12.6,11.4 Hz,H-17),2.28(2H,m,H-23),1.99(2H,m,H-1),1.73(1H,m,H-5),1.69(3H,s,H-26),1.62(3H,s,H-27),1.38(3H,s,H-30),1.12(3H,s,H-28),1.05(3H,s,H-29),1.04(3H,s,H-19),0.83(3H,s,H-18)。13C-NMR(150 MHz,CDCl3)δC:216.1(C-3),180.4(C-21),143.0(C-8),132.7(C-25),123.6(C-24),119.2(C-7),82.1(C-16),72.0(C-12),54.9(C-13),53.2(C-17),52.4(C-5),48.0(C-9),47.8(C-4),45.5(C-20),44.4(C-14),38.4(C-1),36.3(C-15),35.3(C-10),34.7(C-2),33.8(C-30),30.1(C-11),29.1(C-22),26.0(C-23),25.7(C-26),24.4(C-29),24.3(C-6),21.4(C-28),19.9(C-27),17.9(C-18),12.6(C-19)。與文獻(xiàn)[6]比對,鑒定化合物5為12β-hydroxykulactone。

化合物6:白色粉末,分子式C30H46O4,ESI-MSm/z:493[M+Na]+。1H-NMR(600 MHz,CDCl3)δH:5.35(1H,d,J=3.0 Hz,H-7),5.13 (1H,t,J=6.6 Hz,H-24),4.20(1H,ddd,J=11.4,10.3,7.3 Hz,H-16),4.03(2H,dt,J=9.7,4.9 Hz,H-12),2.54(1H,dd,J=12.6,11.4 Hz,H-17),2.47(1H,ddt,J=12.8,6.5,3.0 Hz,H-9),2.40(2H,m,H-20),2.38,1.42(2H,m,H-11),2.29,1.74(2H,dd,J=13.7,10.2 Hz,H-15),2.20,2.13(2H,m,H-23),2.12,1.99(2H,m,H-6),1.96,1.50(2H,m,H-22),1.93,1.62(2H,m,H-2),1.82(1H,dd,J=12.2,5.8 Hz,H-5),1.71(2H,d,J=1.6 Hz,H-26),1.65(3H,d,J=1.3 Hz,H-27),1.59,1.40(2H,m,H-1),1.37(3H,s,H-30),0.97(3H,s,H-28),0.94(3H,s,H-29),0.86(3H,s,H-18),0.82(3H,s,H-19)。13C-NMR(150 MHz,CDCl3)δC:180.6(C-21),143.0(C-8),132.7(C-25),123.7(C-24),119.3(C-7),82.2(C-16),76.0(C-3),72.2(C-12),54.9(C-14),53.2(C-17),48.2(C-9),45.5(C-20),44.6(C-5),44.4(C-13),37.4(C-4),36.2(C-15),35.1(C-10),33.6(C-30),31.1(C-1),30.3(C-11),29.1(C-22),27.7(C-28),26.0(C-23),25.7(C-26),25.3(C-2),23.9(C-6),21.6(C-29),19.8(C-18),17.9(C-27),12.8(C-19)。與文獻(xiàn)[7]比對,鑒定化合物6為mesendanin O。

化合物7:無色油狀,分子式C30H46O4,ESI-MSm/z:491[M+Na]+。1H-NMR(600 MHz,CDCl3)δH:5.48(1H,dd,J=4.2,2.8 Hz,H-7), 5.12(1H,m,H-24),4.50(1H,m,H-6),4.17(1H,ddd,J=11.4,10.1,7.6 Hz,H-16),2.88(2H,td,J=14.1,5.5 Hz,H-2),2.32,1.77(2H,m,H-15),2.00(1H,m,H-17),1.71(3H,d,J=1.4 Hz,H-26),1.64(3H,s,H-27),1.54(3H,s,H-28),1.51(1H,dd,J=4.3,2.6 Hz,H-5),1.33(3H,s,H-30),1.28(3H,s,H-29),1.25(3H,s,H-19),0.94(3H,s,H-18)。13C-NMR(150 MHz,CDCl3)δC:215.8(C-3),180.4(C-21),146.0(C-8),132.8(C-25),123.3(C-24),122.0(C-7),82.1(C-16),67.0(C-6),58.0(C-17),56.7(C-5),55.1(C-14),49.0(C-4),48.8(C-9),45.4(C-20),40.0(C-1),39.4(C-13),35.5(C-10),35.4(C-15),34.6(C-2),31.4(C-30),29.5(C-22),29.2(C-12),26.0(C-23),25.7(C-26),24.7(C-28),23.9(C-29),21.4(C-18),17.9(C-27),16.8(C-11),15.0(C-19)。與文獻(xiàn)[8]比對,鑒定化合物7為6β-hydroxykulactone。

化合物8:無色粉末,分子式C30H48O4,ESI-MSm/z:495[M+Na]+。1H-NMR(600 MHz,CDCl3)δH:5.31(1H,d,J=3.0 Hz,H-7),5.05(2H,brs,H-26),4.30(1H,m,H-24),4.06(1H,m,H-16),2.78,2.21(2H,td,J=14.4,5.4 Hz,H-2),2.27(1H,m,H-9),2.11(2H,ddd,J=11.5,8.9,5.6 Hz,H-6),2.07,2.00(2H,m,H-15),1.92,1.41(2H,m,H-1),1.88,1.56(2H,m,H-12),1.73(3H,s,H-27),1.73,1.39(2H,m,H-23),1.69(1H,m,H-5),1.65(2H,d,J=5.1 Hz,H-11),1.62(1H,m,H-20),1.59,1.08(2H,m,H-22),1.48(1H,m,H-17),1.27(3H,s,H-30),1.14(3H,s,H-28),1.06(3H,s,H-29),1.05(3H,s,H-21),1.04(3H,d,J=1.5 Hz,H-19),0.84(3H,d,J=2.1 Hz,H-18)。13C-NMR(150 MHz,CDCl3)δC:216.9(C-3),145.1(C-8),143.8(C-25),118.2(C-7),114.4(C-26),89.8(C-24),78.1(C-16),62.5(C-17),52.4(C-5),49.9(C-14),47.9(C-4),47.9(C-9),45.8(C-15),45.4(C-13),38.5(C-1),35.1(C-10),34.9(C-2),34.1(C-20),33.2(C-12),30.7(C-22),27.9(C-30),27.8(C-23),24.5(C-28),24.4(C-6),23.5(C-18),21.6(C-29),18.5(C-21),18.2(C-11),17.2(C-27),12.8(C-19)。與文獻(xiàn)[9]比對,鑒定化合物8為meliasenin L。

化合物9:無色晶體,熔點(diǎn)78 ℃～82 ℃,分子式C30H48O4,ESI-MSm/z:479[M+Na]+。1H-NMR(600 MHz,CDCl3)δH:5.29(1H,s,H-7),4.93(2H,dt,J=1.8,0.9 Hz,H-26),4.85(1H,t,J=1.7 Hz,H-16),4.03(1H,d,J=5.3 Hz,H-24),2.75,2.24(2H,m,H-2),2.25(1H,m,H-9),2.11,1.48(2H,m,H-15),2.10(2H,m,H-6),1.99,1.44(2H,m,H-1),1.90,1.60(2H,m,H-12),1.73(3H,m,H-27),1.71(1H,d,J=3.4 Hz,H-5),1.69,1.45(2H,m,H-23),1.65,0.98(2H,m,H-22),1.62(1H,m,H-20),1.57(2H,m,H-11),1.44(1H,m,H-17),1.25(3H,s,H-30),1.11(3H,s,H-29),1.05(3H,d,J=1.6 Hz,H-21),1.04(3H,s,H-28),1.01(3H,s,H-19),0.82(3H,s,H-18)。13C-NMR(150 MHz,CDCl3)δC:216.8(C-3),147.7(C-25),145.1(C-8),118.1(C-7),111.1(C-26),78.0(C-16),76.5(C-24),62.5(C-17),52.4(C-5),49.9(C-14),47.9(C-4),47.9(C-9),45.7(C-15),45.4(C-13),38.4(C-1),35.0(C-10),34.9(C-2),34.2(C-20),33.2(C-12),31.8(C-23),30.8(C-22),27.8(C-30),24.5(C-28),24.3(C-6),23.5(C-18),21.6(C-29),18.6(C-21),18.2(C-11),17.4(C-27),12.8(C-19)。與文獻(xiàn)[8]比對,鑒定化合物9為meliastatin 5。

化合物10:白色粉末,分子式C30H48O5,ESI-MSm/z:511[M+Na]+。1H-NMR(600 MHz,CDCl3)δH:5.32(1H,m,H-7),4.17(1H,dd,J=12.0,3.5 Hz,H-24),4.02(1H,dd,J=8.5,4.9 Hz,H-16),3.50(1H,dd,J=6.8,4.0 Hz,H-3),2.66(1H,td,J=11.2,7.9 Hz,H-20),2.46,1.97(2H,m,H-2),2.30(1H,m,H-9),2.09(1H,m,H-17),1.91(2H,m,H-6),1.83(2H,m,H-12),1.90,1.39(2H,m,H-22),1.79(2H,dd,J=12.1,5.7 Hz,H-23),1.75(1H,d,J=1.0 Hz,H-5),1.73(2H,d,J=1.0 Hz,H-15),1.64,1.56(2H,m,H-11),1.53(2H,m,H-1),1.33(3H,s,H-26),1.31(3H,s,H-30),1.26(3H,s,H-27),0.96(3H,s,H-18),0.94(3H,s,H-28),0.93(3H,s,H-29),0.81(3H,s,H-19)。13C-NMR(150 MHz,CDCl3)δC:178.5(C-21),144.6 (C-8),118.8(C-7),83.4(C-24),77.6(C-16),76.3(C-3),71.1(C-25),57.9(C-17),50.0(C-14),48.1(C-9),45.8(C-13),44.6(C-15),43.9(C-5),41.9(C-20),37.4(C-4),34.8(C-10),33.6(C-12),31.2(C-1),27.8(C-28),27.3(C-30),26.2(C-26),25.4(C-2),24.4(C-27),23.9(C-23),23.2(C-18),22.7(C-22),21.8(C-29),21.3(C-6),17.9(C-11),13.0(C-19)。與文獻(xiàn)[10]比對,鑒定化合物10為toosendine G。

以上10種化合物均為甘遂烷型三萜類化合物,其中化合物2和5首次從川楝中分離得到。