羅 彭,鄧祖帥,王佳佳,沈章陽(yáng),潘為高

(廣西中醫(yī)藥大學(xué),廣西南寧 530200)

線葉報(bào)春苣苔[Primulinalinearifolia(W.T.Wang)Yin Z.Wang]為苦苣苔科(Gesneriaceae)報(bào)春苣苔屬(Primulina)多年生草本,壯藥名為接骼紅,中藥名為接骨草,廣西地域性壯族藥用植物,主要分布于中國(guó)廣西南部,例如南寧市邕寧區(qū)、大新縣、隆安縣、武鳴區(qū)[1];其根狀莖可供藥用,有清熱解毒之效,用于調(diào)氣道、續(xù)筋骨、散瘀腫,治療勞傷咳嗽、骨折、跌打腫痛等病癥[1,2]。

本課題組前期報(bào)道了線葉報(bào)春苣苔含有多種醌類成分(蒽醌、α-董尼酮、萘駢董尼酮),并具有優(yōu)良抗腫瘤作用[3],但關(guān)于該植物的其他成分,國(guó)內(nèi)外目前尚無(wú)研究報(bào)道,僅有植物生理方面的報(bào)道[4]。其同屬植物的化學(xué)成分或藥理研究方面,螞蝗七[P.fimbrisepala(Hand.-Mazz.)Yin Z.Wang]含有黃酮和甾醇苷類[5];牛耳朵[P.eburnea(Hance)Yin Z.Wang]含有蒽醌和苯乙醇苷類,其苯乙醇苷類具有抗癌活性[6-8];報(bào)春苣苔(P.tabacumHance)含有的阿克苷具有抗腫瘤及抗腫瘤轉(zhuǎn)移、抗凋亡作用[9,10];紅藥[P.longgangensis(W.T.Wang) Yan Liu &Yin Z.Wang]含蒽醌、香草酸、車前草苷、三萜及苷類等[11-13]。

除了上述典籍記載的藥用價(jià)值[1,2],根據(jù)歷次廣西植物資源普查科考研究得知,在桂南馬山、武鳴、隆安等區(qū)縣,民間也稱線葉報(bào)春苣苔為紅接骨草、吊石牛,全草用于勞傷咳嗽、崩漏、肝炎,外治跌打損傷、骨折、消腫,是壯族民間赤腳醫(yī)生的常用藥。但該藥用植物除其所含醌類被報(bào)道外[4],其他化學(xué)成分未見有相關(guān)研究,藥效物質(zhì)基礎(chǔ)不明,藥材質(zhì)量標(biāo)準(zhǔn)尚未制定。本研究較系統(tǒng)地對(duì)其化學(xué)成分進(jìn)行研究,擬為該藥用植物傳統(tǒng)藥效的物質(zhì)基礎(chǔ)研究、質(zhì)量標(biāo)準(zhǔn)制定和開發(fā)應(yīng)用提供借鑒。

1 材料與方法

1.1 植物藥材

線葉報(bào)春苣苔藥材采集于廣西南寧市隆安縣,生長(zhǎng)于該縣山地的石山蔭處及疏林中石上,經(jīng)廣西中醫(yī)藥大學(xué)朱意麟實(shí)驗(yàn)師和韋松基教授鑒定為苦苣苔科報(bào)春苣苔屬線葉報(bào)春苣苔[Primulinalinearifolia(W.T.Wang)Yin Z.Wang]。藥材全株,陰干、粉碎,于4 ℃密閉保存?zhèn)溆谩?/p>

1.2 儀器與試劑

Waters Auto Spec Premier P776質(zhì)譜儀(Waters,美國(guó));Bruker Avance-500 型和Bruker AM-800型核磁共振波譜儀(Bruker,瑞士);ZF-2三用紫外儀(上海市安亭電子儀器廠);XT4-100A顯微熔點(diǎn)測(cè)定儀(北京市科儀電光儀器廠)。

柱層析硅膠GF254(200-300目、300-400目)和薄層用硅膠G板(青島海洋化工有限公司);D101大孔吸附樹脂(上海展云化工有限公司);聚酰胺柱色譜填料(80-120目,100-200目,常州市長(zhǎng)豐化工有限公司);Sephadex LH-20(法瑪西亞公司,瑞典);ODS柱色譜填料(YMC公司,日本);MCI(三菱化學(xué)公司,日本);乙醇、石油醚、乙酸乙酯、二氯甲烷、三氯甲烷、丙酮、甲醇(分析純,成都市科隆化學(xué)品有限公司)。

1.3 方法

線葉報(bào)春苣苔全草粗粉8.65 kg,用15倍量85%乙醇,在60 ℃條件下熱攪拌,重復(fù)提取3次,每次12 h;濾渣在相同條件下,用15倍量50%乙醇重復(fù)提取3次,每次12 h。合并提取液,減壓濃縮,濃縮液水浴揮干至無(wú)醇味,并加適量水混懸,依次用石油醚(沸程60-90 ℃)、乙酸乙酯、正丁醇萃取,將萃取液分別減壓濃縮,得石油醚部位(140.0 g)、乙酸乙酯部位(160.0 g)、正丁醇部位(205.0 g)、水部位(203.0 g)。

取石油醚部位130.0 g,經(jīng)硅膠柱層析(100-200目),用石油醚-乙酸乙酯(100∶1→50∶1→25∶1→15∶1→8∶1→5∶1→3∶1→2∶1→1∶1→0∶1,V/V)梯度洗脫,得20個(gè)流分;各流分分別經(jīng)硅膠柱色譜(200-300目,流動(dòng)相為石油醚-乙酸乙酯或石油醚-丙酮)反復(fù)分離,并結(jié)合Sephadex LH-20柱色譜(三氯甲烷∶甲醇=1∶1,V/V)和重結(jié)晶進(jìn)行純化,最終獲得10個(gè)化合物:1(5 mg)、2(4 mg)、3(300 mg)、4(5 mg)、5(3 mg)、6(3 mg)、7(3 mg)、8(5 mg)、9(150 mg)、10(30 mg)。

取乙酸乙酯部位130.0 g,用硅膠柱色譜(100-200目)分離,先以石油醚-乙酸乙酯(100∶1→15∶1,V/V)洗脫,接著用三氯甲烷-甲醇(100∶1→25∶1→15∶1→8∶1→5∶1→3∶1→2∶1→1∶1→0∶1,V/V)洗脫;各流分經(jīng)硅膠柱色譜(200-300目,流動(dòng)相為石油醚-二氯甲烷)反復(fù)分離,并結(jié)合Sephadex LH-20柱色譜(三氯甲烷∶甲醇=1∶1,V/V)和重結(jié)晶進(jìn)行純化,最終獲得4個(gè)化合物:11(7 mg)、12(6 mg)、13(9 mg)、14(6 mg)。

取正丁醇部位150.0 g,用大孔樹脂(D101)柱色譜進(jìn)行分離,采用水→30%乙醇→50%乙醇→70%乙醇→無(wú)水乙醇梯度洗脫;所得各流分,根據(jù)極性等特性差異,分別用聚酰胺柱色譜(100-200目,水→30%乙醇→50%乙醇→70%乙醇→無(wú)水乙醇洗脫)或硅膠柱色譜(200-300目,三氯甲烷-甲醇:20∶1→10∶1→0∶1,V/V)或反相ODS柱色譜(水→30%乙醇→50%乙醇→70%乙醇→100%乙醇)進(jìn)一步分離,最終獲得3個(gè)化合物:15(7 mg)、16(9 mg)、17(8 mg)。

2 結(jié)果與分析

化合物1:無(wú)色粉末,m.p.87-89 ℃。EI-MS (m/z):410[M]+。1H-NMR (500 MHz,CDCl3)δ:0.88 (3H,t,J=7.1 Hz,H-27),1.25 (46H,m,H-4-26),1.63 (2H,m,H-3),2.35 (2H,t,J=7.5 Hz,H-2);13C-NMR and DEPT (125 MHz,CDCl3)δ:14.1 (CH3,C-27),22.7 (CH2,C-26),24.7 (CH2,C-3),29.1-29.7 (CH2,C-4-24),31.9 (CH2,C-25),34.1 (CH2,C-2),180.3 (qC,C-1)。以上數(shù)據(jù)與文獻(xiàn)[14]報(bào)道基本一致,化合物1鑒定為二十七酸(Heptacosanoic acid,C27H54O2)。

化合物2:白色粉末,m.p.52-53 ℃。EI-MS (m/z): 412 [M]+。1H-NMR (500 MHz,CDCl3)δ:5.35 (2H,m,H-13,14),4.21 (1H,dd,J=11.5,5.0 Hz,H-1′a),4.15 (1H,dd,J=11.5,5 Hz,H-1′b),3.93 (1H,m,H-2′),3.70 (1H,dd,J=11.5,6.0 Hz,H-3′a),3.60 (1H,dd,J=11.5,6.0 Hz,H-3′b),2.35 (2H,t,J=7.6 Hz,H-2),2.20 (2H,t,J=7.5 Hz,H-15),2.00 (2H,m,H-12),1.63 (2H,m,H-3),1.26 (28H,m,H-4-11,H-16-21),0.88 (3H,t,J=7.5 Hz,H-22);13C-NMR and DEPT (125 MHz,CDCl3)δ:174.4 (qC,C-1),129.9 (CH,C-13),129.9 (CH,C-14),70.2 (CH,C-2′),65.2 (CH2,C-1′),63.3 (CH2,C-3′),35.9 (CH2,C-2),34.1 (CH2,C-12,15),31.9 (CH2,C-20),29.8,29.6,29.6,29.5,29.5,29.4,29.4,29.3,29.2,29.1,27.2,25.5 (CH2,C-4-11,C-16-19),24.9 (CH2,C-3),22.7 (CH2,C-21),14.1 (CH3,C-22)。對(duì)照文獻(xiàn)[15]的核磁數(shù)據(jù),化合物2鑒定為芥酸單甘油酯(Monoerucin,C25H48O4)。

化合物3:無(wú)色針狀結(jié)晶,m.p.141-142 ℃。EI-MS (m/z):414 [M]+。1H-NMR (500 MHz,CDCl3)δ:5.35 (1H,m,H-6),3.52 (1H,m,H-3);13C-NMR and DEPT (125 MHz,CDCl3)δ:140.5 (qC,C-5),121.9 (CH,C-6),71.9 (CH,C-3),56.8 (CH,C-14),56.1 (CH,C-17),50.3 (CH,C-9),46.0 (CH,C-24),42.2 (qC,C-13),40.0 (CH2,C-4),39.7 (CH2,C-12),37.3 (CH2,C-1),36.6 (qC,C-10),36.2 (CH,C-20),34.0 (CH2,C-22),31.8 (CH2,C-7),31.8 (CH,C-8),31.1 (CH2,C-2),29.3 (CH,C-25),28.4 (CH2,C-16),26.2 (CH2,C-23),24.5 (CH2,C-15),23.1 (CH2,C-28),21.3 (CH2,C-11),20.1 (CH3,C-27),19.5 (CH3,C-19),19.0 (CH3,C-26),18.9 (CH3,C-21),12.3 (CH3,C-29),12.0 (CH3,C-18)。對(duì)照文獻(xiàn)[16]的核磁數(shù)據(jù),化合物3鑒定為β-谷甾醇(β-Sitosterol,C29H50O)。

化合物4:無(wú)色針狀結(jié)晶,易溶于三氯甲烷,m.p.85-87 ℃。EI-MS (m/z): 412[M]+。1H-NMR (500 MHz,CDCl3)δ: 5.72 (1H,s,H-4),2.40 (2H,m,H-2);13C-NMR and DEPT (125 MHz,CDCl3)δ:11.9 (CH3,C-18),11.9 (CH3,C-29),17.4 (CH3,C-26),18.7 (CH3,C-21),19.0 (CH3,C-19),19.8 (CH3,C-27),21.0 (CH2,C-11),23.0 (CH2,C-28),24.2 (CH2,C-15),26.0 (CH,C-25),28.2 (CH2,C-16),29.1 (CH2,C-23),32.0 (CH2,C-2),32.9 (CH2,C-6),33.8 (CH2,C-7),34.0 (CH2,C-22),35.6 (CH2,C-1),35.7 (CH,C-8),36.1 (CH,C-20),38.6 (qC,C-10),39.6 (CH2,C-12),42.4 (qC,C-13),45.8 (CH,C-24),53.8 (CH,C-9),55.8 (CH,C-17),56.0 (CH,C-14),123.7 (CH,C-4),171.7 (qC,C-5),199.7 (qC,C-3)。以上數(shù)據(jù)與文獻(xiàn)[17]報(bào)道基本一致,化合物4鑒定為β-扶桑甾醇氧化物(β-sitostenone,C29H48O)。

化合物5:無(wú)色片狀結(jié)晶,m.p.76-77 ℃。 EI-MS (m/z):586 [M]+。1H-NMR (800 MHz,CDCl3)δ:7.08 (2H,d,J=8.4 Hz,H-2,6),6.76 (2H,J= 8.4 Hz,H-3,5),4.23 (2H,t,J=7.1 Hz,H-8),2.86 (2H,t,J=7.1 Hz,H-7),2.27 (2H,t,J=7.6 Hz,H-2′),1.58 (2H,m,H-3′),1.25 (64H,m,H-4′-30′),0.88 (3H,t,J=7.1 Hz,H-31′);13C-NMR and DEPT (200 MHz,CDCl3)δ:173.9 (qC,C-1′),154.2 (qC,C-4),130.1 (CH,C-2,6),129.9 (qC,C-1),115.3 (CH,C-3,5),64.9 (C-8),34.3 (C-7),34.3 (C-2′),31.9 (C-29′),29.1-29.7 (C-4′-28′),24.9 (C-3′),22.7 (C-30′),14.1 (CH3,C-31′)。對(duì)照文獻(xiàn)[18]的Heitziethanoid A光譜數(shù)據(jù),化合物5結(jié)構(gòu)鑒定為三十一烷酸對(duì)羥基苯乙酯[Hentriacontanoic acid,2- (4-hydroxyphenyl)ethyl ester,C39H70O3]。

化合物6:無(wú)色簇狀結(jié)晶,m.p.210-212 ℃。EI-MS (m/z):168 [M]+。1H-NMR (800 MHz,CD3OD)δ:7.51 (1H,d,J=1.9 Hz,H-2),7.55 (1H,dd,J=8.3,1.9 Hz,H-6),6.83 (1H,d,J=8.3 Hz,H-5),3.87 (3H,s,OCH3);13C-NMR and DEPT (200 MHz,CD3OD)δ:169.5 (qC,C-7),151.3 (qC,C-4),147.5 (qC,C-3),124.6 (CH,C-6),122.3 (qC,C-1),115.0 (CH,C-5),113.0 (CH,C-2),56.1 (OCH3)。以上數(shù)據(jù)與文獻(xiàn)[19]報(bào)道基本一致,故化合物6鑒定為香草酸(Vanillic acid,C8H8O4)。

化合物7:無(wú)色片狀結(jié)晶,m.p.290-291 ℃。EI-MS (m/z): 456[M]+。1H-NMR (800 MHz,CDCl3)δ: 5.25 (1H,t,J=3.8 Hz,H-12),3.22 (1H,dd,J=11.5,4.6Hz,H-3);13C-NMR and DEPT (200 MHz,CDCl3)δ:181.8 (qC,C-28),137.9 (qC,C-13),125.8 (CH,C-12),79.0 (CH,C-3),55.2 (CH,C-5),52.6 (CH,C-18),47.9 (qC,C-17),47.5 (CH,C-9),42.0 (qC,C-14),39.5 (qC,C-10),39.0 (CH,C-20),38.8 (CH,C-19),38.7 (qC,C-4),38.6 (CH2,C-22),37.0 (qC,C-8),36.7 (CH2,C-1),32.9 (CH2,C-7),30.6 (CH2,C-21),28.1 (CH3,C-23),28.0 (CH2,C-2),27.2 (CH2,C-15),24.1 (CH2,C-16),23.6 (CH3,C-27),23.3 (CH2,C-11),21.2 (CH3,C-30),18.3 (CH2,C-6),17.0 (CH3,C-26),17.0 (CH3,C-29),15.6 (CH3,C-24),15.5 (CH3,C-25)?；衔?光譜數(shù)據(jù)與熊果酸數(shù)據(jù)[20]基本一致,故確定化合物7為熊果酸(Ursolic acid,C30H48O3)。

化合物8:白色粉末,m.p.92-94 ℃。EI-MS (m/z): 498 [M]+。1H-NMR (800 MHz,CD3OD)δ:4.05 (2H,m,H-1′),3.81 (1H,m,H-2′),3.55 (1H,dd,J=11.5,4.2 Hz,H-3′a),3.48 (1H,dd,J=11.5,6.2 Hz,H-3′b),2.28 (2H,t,J=7.6 Hz,H-2),1.55 (2H,t,J=7.6 Hz,H-3),1.14-1.38 (48H,m,H-4-27),0.81 (3H,t,J=7.0 Hz,H-28);13C-NMR and DEPT (200 MHz,CD3OD)δ:174.6 (qC,C-1),70.0 (CH,C-2′),65.2 (CH2,C-1′),63.2 (CH2,C-3′),34.9 (CH2,C-2),31.9 (CH2,C-26),29.1-29.7 (CH2,C4-25),24.9 (CH2,C-3),22.7 (CH2,C-27),14.0 (CH3,C-28)。對(duì)照文獻(xiàn)[21]中的二十二(或二十四)酸甘油酯波譜數(shù)據(jù)進(jìn)行歸屬,化合物8鑒定為1-二十八酸甘油酯(1-Octacosanoyl glyceride,C31H62O4)。

化合物9:白色粉末,m.p.301-303 ℃,易溶于三氯甲烷、甲醇。EI-MS (m/z):398 [M-C6H11O6]+。1H-NMR(500 MHz,Pyridine-d6)δ:5.33 (1H,m,H-6),4.36 (1H,d,J=7.9 Hz,H-1′),3.79-3.81 (1H,m,H-3′),3.68-3.70 (1H,m,H-4′),3.53-3.57 (1H,m,H-3),3.35-3.37 (2H,m,H-6′),3.23-3.24 (1H,m,H-5′),3.16-3.19 (1H,m,H-2′);13C-NMR and DEPT (125 MHz,Pyridine-d6)δ:140.7 (qC,C-5),122.4 (CH,C-6),101.5 (CH,C-1′),79.5 (CH,C-3),76.9 (CH,C-3′),76.3 (CH,C-5′),74.0 (CH,C-2′),70.6 (CH,C-4′),62.1 (CH2,C-6′),57.1 (CH,C-17),56.4 (CH,C-14),50.6 (CH,C-9),46.2 (CH,C-24),42.7 (qC,C-13),40.1 (CH2,C-12),39.0 (CH2,C-4),37.6 (CH2,C-1),37.1 (qC,C-10),36.5 (CH,C-20),34.3 (CH2,C-22),32.3 (CH2,C-7),32.3 (CH,C-8),29.9 (CH2,C-25),29.5 (CH,C-16),28.6 (CH2,C-2),26.4 (CH2,C-23),24.6 (CH2,C-15),23.4 (CH2,C-28),21.4 (CH2,C-11),20.0 (CH3,C-21),19.6 (CH3,C-26),19.2 (CH3,C-19),19.0 (CH3,C-27),12.2 (CH3,C-18),12.1 (CH3,C-29)。以上數(shù)據(jù)與文獻(xiàn)[22,23]報(bào)道基本一致,故化合物9鑒定為胡蘿卜苷(Daucosterol,C35H60O6)。

化合物10:灰白色固體,m.p.187-189 ℃。ESI-MS (m/z):477[M-H]-。1H-NMR (800 MHz,CD3OD)δ:6.70 (1H,d,J=2.0 Hz,H-2),6.69 (1H,d,J=8.0 Hz,H-5),6.53 (1H,dd,J=8.0,2.0 Hz,H-6),3.62 (2H,m,H-8),2.77 (2H,m,H-7)(phenethyl alcohol moiety);7.56 (1H,d,J=15.8 Hz,H-7′),7.01 (1H,d,J=2.0 Hz,H-2′),6.88 (1H,dd,J=8.0,2.0 Hz,H-6′),6.76 (1H,d,J=8.0 Hz,H-5′),6.22 (1H,d,J=15.8 Hz,H-8′)(caffeic acid moiety);4.83 (1H,t,J=9.6 Hz,H-4″),4.31 (1H,d,J=7.8 Hz,H-1″),3.70 (1H,m,H-3″),3.54 (1H,m,H-5″),3.43 (1H,m,H-2″),3.31 (2H,m,H-6″)(glucose moiety)。13C-NMR and DEPT (200 MHz,CD3OD)δ:144.9 (qC,C-3),143.5 (qC,C-4),130.7 (qC,C-1),120.7 (CH,C-6),116.3 (CH,C-2),115.6 (CH,C-5) ,71.4 (CH2,C-8),35.7 (CH2,C-7)(phenethyl alcohol moiety);168.2 (qC,C-9′),148.6 (qC,C-4′),147.3 (CH,C-7′),145.6 (qC,C-3′),126.8 (qC,C-1′),122.6 (CH,C-6′),115.8 (CH,C-5′),114.6 (CH,C-2′),113.7 (CH,C-8′)(caffeic acid moiety);103.3 (CH,C-1″),78.0 (CH,C-4″),75.1 (CH,C-2″),74.7 (CH,C-5″),74.3 (CH,C-3″),61.6 (CH2,C-6″)(glucose moiety)。對(duì)照文獻(xiàn)[10,24]的核磁數(shù)據(jù),化合物10鑒定為木通苯乙醇苷A(Calceolarioside A,C23H26O11)。

化合物11:白色粉末,溶于甲醇,鹽酸-鎂粉反應(yīng)呈陽(yáng)性,m.p.254-255 ℃。EI-MS (m/z): 450[M]+。1H-NMR (500 MHz,CD3OD)δ:6.89(1H,s,H-2′),6.76(2H,s,H-5′,6′),5.95(1H,s,H-8),5.30 (1H,dd,J=12.3,3.0 Hz,H-2),4.78(1H,d,J=9.9 Hz,H-1″),4.10(1H,m,H-2″),3.85 (1H,d,J=12.0 Hz,H-6″),3.71 (1H,dd,J=12.0,5.4 Hz,H-6″),3.35-3.43 (3H,m,H-3″,4″,5″),3.10(1H,dd,J=17.2,12.3 Hz,H-3),2.75 (1H,dd,J=17.2,3.0 Hz,H-3);13C-NMR and DEPT (125 MHz,CD3OD)δ:198.1 (qC,C-4),167.3 (qC,C-7),164.3 (qC,C-5),164.2 (qC,C-9),146.9 (qC,C-4′),146.5 (qC,C-3′),131.6 (qC,C-1′),119.2 (CH,C-6′),116.2 (CH,C-5′),114.7 (CH,C-2′),105.9 (qC,C-6),103.2 (qC,C-10),96.3 (CH,C-8),82.5 (CH,C-5″),80.5 (CH,C-3″),80.2 (CH,C-2),75.2 (CH,C-1″),72.6 (CH,C-2″),71.8 (CH,C-4″),62.9 (CH2,C-6″) ,44.0 (CH2,C-3)?；衔?1光譜數(shù)據(jù)與文獻(xiàn)中5,7,3′,4′-四羥基-6-C-β-D-葡萄糖二氫黃酮碳苷的數(shù)據(jù)[25]一致,故鑒定其為5,7,3′,4′-四羥基-6-C-β-D-葡萄糖二氫黃酮碳苷(5,7,3′,4′-tetrahydroxy -6-C-β-D- glucosyl-dihydroflavonol,C21H22O11)。

化合物12:灰白色固體,易吸潮,m.p.136-138 ℃。經(jīng)FeCl3-K3[Fe (CN)6]試劑鑒別,顯藍(lán)色,可能為酚類物質(zhì)。ESI-MS (m/z):639 [M-H]-。1H-NMR (500 MHz,CD3OD)δ:6.71 (1H,d,J=2.0 Hz,H-2),6.69 (1H,d,J=8.0 Hz,H-5),6.57 (1H,dd,J=8.0,2.0 Hz,H-6),3.66 (2H,m,H-8),2.79 (2H,m,H-7)(phenethyl alcohol moiety);7.60 (1H,d,J=15.9 Hz,H-7′),7.09 (1H,d,J=2.0 Hz,H-2′),6.99 (1H,dd,J=8.0,2.0 Hz,H-6′),6.80 (1H,d,J=8.0 Hz,H-5′),6.34 (1H,d,J=15.9 Hz,H-8′)(caffeic acid moiety);4.41 (1H,t,J=7.9 Hz,H-1″),4.03 (1H,m,H-4″),3.94 (1H,t,J=9.3Hz,H-3″),3.50 (2H,m,H-6″),3.25 (1H,m,H-5″),3.16 (1H,m,H-2″)(glucose moiety 1);4.55 (1H,t,J=7.8 Hz,H-1?),3.74 (1H,m,H-4?),3.72 (1H,m,H-3?),3.54 (2H,m,H-6?),3.32 (1H,m,H-5?),3.19 (1H,m,H-2?)(glucose moiety 2)。13C-NMR and DEPT (125 MHz,CD3OD)δ:146.1 (qC,C-3),144.6 (qC,C-4),131.4 (qC,C-1),121.4 (CH,C-6),117.2 (CH,C-2),116.4 (CH,C-5) ,72.2 (CH2,C-8),36.5 (CH2,C-7)(phenethyl alcohol moiety);168.6 (qC,C-9′),149.7 (qC,C-4′),147.5 (CH,C-7′),146.8 (qC,C-3′),127.7 (qC,C-1′),123.2 (CH,C-6′),116.6 (CH,C-5′),115.3 (CH,C-2′),115.2 (CH,C-8′)(caffeic acid moiety);103.9 (CH,C-1″),84.3 (CH,C-3″),76.0 (CH,C-2″),75.8 (CH,C-5″),70.9 (CH,C-4″),62.3 (CH2,C-6″)(glucose moiety 1);105.8 (CH,C-1?),77.8 (CH,C-3?),77.6 (CH,C-5?),75.0 (CH,C-2?),71.1 (CH,C-4?),62.4 (CH2,C-6?)(glucose moiety 2)。對(duì)照文獻(xiàn)[26,27]的核磁數(shù)據(jù),確定化合物12為大車前苷(Plantamajoside,C29H36O16)。

化合物13:白色粉末,10% H2SO4顯紫色,易溶于三氯甲烷、甲醇、乙醇,m.p.278-280 ℃。ESI-MS (m/z):575 [M+H]+。1H-NMR (500 MHz,CDCl3+CD3OD)δ:5.07-5.16 (1H,m,H-22,23),5.02 (1H,m,H-7),5.00 (1H,d,J=7.6 Hz,H-1′),4.01 (1H,m,H-3);13C-NMR and DEPT (125 MHz,CDCl3+CD3OD)δ:139.5 (qC,C-8),138.4 (CH,C-22),129.8 (CH,C-23),117.7 (CH,C-7),102.2 (CH,C-1′),78.4 (CH,C-3′),78.0 (CH,C-5′),77.5 (CH,C-3),75.2 (CH,C-2′),71.9 (CH,C-4′),63.0 (CH2,C-6′),56.3 (CH,C-17),55.3 (CH,C-14),51.4 (CH,C-24),49.7 (CH,C-9),43.5 (qC,C-13),40.5 (CH,C-5),40.3(CH,C-20),39.7 (CH2,C-12),37.4 (CH2,C-1),34.7 (qC,C-10),34.6 (CH2,C-4),32.1 (CH,C-25),30.1 (CH2,C-2,6),28.6 (CH2,C-16),25.5 (CH2,C-28),23.3 (CH2,C-15),21.7 (CH2,C-11),21.3 (CH3,C-27),21.2 (CH3,C-21),19.3 (CH3,C-26),12.8 (CH3,C-19),12.2 (CH3,C-29),12.1(CH3,C-18)。對(duì)照文獻(xiàn)[28]的核磁數(shù)據(jù),化合物13鑒定為α-菠甾醇-3-O-β-D-葡萄糖苷(α-spinasteryl-3-O-β-D-glucoside,C35H58O6)。

化合物14:白色粉末,易溶于三氯甲烷、甲醇,m.p.338 ℃。EI-MS (m/z): 226 [M]+。1H-NMR (500 MHz,CDCl3)δ:7.31 (1H,s,NH,H-1),7.28 (1H,s,NH,H-1),3.05 (4H,m,H-7,14),2.05 (4H,m,H-3,10),1.49 (4H,m,H-4,11),1.38 (4H,t,H-6,13),1.19 (4H,t,H-5,12);13C-NMR (125 MHz,CDCl3)δ:174.4 (qC,C-2,9),39.0 (CH2,C-7,14),35.9 (CH2,C-3,10),28.7 (CH2,C-6,13),26.0 (CH2,C-5,12),25.2 (CH2,C-4,11)。對(duì)照文獻(xiàn)[29]的核磁數(shù)據(jù),化合物14鑒定為1,8-二氮雜環(huán)十四烷-2,9-二酮(1,8-diazacyclotetradecane-2,9-dinoe,C12H22N2O2)。

化合物15:無(wú)色油狀物。EI-MS (m/z):506[M]+。1H-NMR (500 MHz,C5D5N)δ:5.45 (2H,m,H-24,25),2.43 (2H,m,H-2),2.06 (2H,m,H-23),1.83 (2H,m,H-26),1.23 (54H,m,H-3-22,H-27-33),0.83 (3H,m,H-34);13C-NMR (125 MHz,C5D5N)δ: 180.1 (qC,C-1),130.2 (CH,C-24,25),36.3 (CH2,C-2),32.1 (CH2,C-23,26),27.5-30.1 (CH2,m,C-4-22,C-27-32),26.3 (CH2,C-3),23.0 (CH2,C-33),14.3 (CH3,C-34)。與文獻(xiàn)[30]中的n-Tetratriacont-24-enoic acid光譜數(shù)據(jù)基本吻合,故化合物15鑒定為24-三十四碳烯酸(n-Tetratriacont-24-enoic acid,C34H66O2)。

化合物16:白色粉末,m.p.110 ℃。EI-MS (m/z):508 [M]+。1H-NMR (500 MHz,C5D5N)δ:2.22 (2H,m,H-2),1.63 (2H,m,H-3),1.25 (60H,m,H-4-33),0.88 (3H,m,H-34);13C-NMR (125 MHz,C5D5N)δ:180.1 (qC,C-1),35.9 (CH2,C-2),31.9 (CH2,C-32),29.3-29.7 (CH2,m,C-4-31),25.7 (CH2,C-3),22.7 (CH2,C-33),14.1 (CH3,C-34)?；衔?6鑒定為正三十四烷酸(n-Tetratriacontanoic acid,C34H68O2)[31]。

化合物17:白色片狀結(jié)晶,m.p.83 ℃。 EI-MS (m/z):590 [M]+。1H-NMR (500 MHz,CDCl3)δ:1.25 (80H,m,H-2-41),0.88 (6H,t,J=7.0 Hz,H-1,42);13C-NMR (125 MHz,CDCl3)δ:31.9 (CH2,C-3,40),29.3-29.7 (CH2,m,C-4-39),22.7 (CH2,C-2,41),14.1 (CH3,C-1,42)。化合物17鑒定為正四十二烷 (Alkane C42,n-C42H86)[32]。

化合物1-17的結(jié)構(gòu)式如圖1所示。

3 討論

三萜或甾類成分最顯著的生理活性為抗炎、保肝等。線葉報(bào)春苣苔所含的1個(gè)五環(huán)三萜和4個(gè)甾體類(包括苷),很可能是其具有清熱解毒、治跌打損傷、散瘀腫(消腫)、治療肝炎等藥效的重要物質(zhì)基礎(chǔ)。本研究發(fā)現(xiàn)線葉報(bào)春苣苔含有2個(gè)苯乙醇-咖啡酰糖苷,該類成分為阿克替苷的同系物,阿克替苷具有神經(jīng)和組織保護(hù)、抗氧化、抗衰老、保肝等多種生理活性,是肉蓯蓉、藏波羅花中的重要滋補(bǔ)成分[33],這與線葉報(bào)春苣苔用于跌打損傷治療的記載相吻合。線葉報(bào)春苣苔含有的二氫黃酮苷類成分亦具有多種生理活性。另外,課題組前期從線葉報(bào)春苣苔中亦分離獲得了系列蒽醌、α-董尼酮和萘駢董尼酮類成分,并報(bào)道了其優(yōu)良抗癌特性[3]。醌類成分具有顯著的抗菌(抗感染)、止血等作用,這與線葉報(bào)春苣苔具有清熱解毒、治勞傷咳嗽、治崩漏的藥效記載相吻合?？傊?化學(xué)成分研究為該植物傳統(tǒng)藥效作用的物質(zhì)基礎(chǔ)研究、質(zhì)量標(biāo)準(zhǔn)制定及深度開發(fā)應(yīng)用提供了重要借鑒。

4 結(jié)論

本研究從線葉報(bào)春苣苔中獲得17個(gè)化合物,按類別可分為飽和或不飽和脂肪酸(1、15、16)、脂肪酸單甘油酯(2、8)、甾體或其苷類(3、4、9、13)、苯乙醇脂肪酸酯(5)、苯甲酸類(6)、五環(huán)三萜(7)、苯乙醇-咖啡酰糖苷(10、12)、二氫黃酮碳苷(11)、環(huán)雙內(nèi)酰胺(14)、飽和烷烴(17),其中多數(shù)是生物活性功能性成分,該研究為廣西地域特色壯藥植物線葉報(bào)春苣苔的質(zhì)量標(biāo)準(zhǔn)研究和深度開發(fā)應(yīng)用提供了良好借鑒。