山螺化學(xué)成分的分離與鑒定

山螺化學(xué)成分的分離與鑒定

山海瑞是桔?？频囊环N廣泛分布于中國(guó)東北、華北、華東和以南的地區(qū)的植物。根藥，包括腫脹、解毒、排膿、加速牧場(chǎng)、抗疲勞等功效。為了開發(fā)山海螺的藥用價(jià)值,筆者對(duì)山海螺的化學(xué)成分作了較為系統(tǒng)的研究。從山海螺干燥根中分離鑒定出11個(gè)化合物,經(jīng)波譜解析和物理常數(shù)測(cè)定為蒲公英萜酮(Ⅰ)、蒲公英萜醇(Ⅱ)、α-菠甾醇(Ⅲ)、順丁烯二酸(Ⅳ)、二十六烷酸甲酯(Ⅴa)、正二十九烷(Ⅴb)、二十四碳酸二十一烷醇酯(Ⅵa)、四十四烷酸甲酯(Ⅵb)、α-菠甾醇-β--葡萄糖苷(Ⅶ)、△7-豆甾烯醇-β--葡萄糖苷(Ⅶb)、豆甾醇-β-D-葡萄糖苷(Ⅶc)。其中Ⅴa、Ⅴb、Ⅵa、Ⅵb、Ⅶa、Ⅶb為首次從該植物中分得。1儀器、試藥和儀器XT-4A型熔點(diǎn)測(cè)定儀(溫度未校正),Cintra-10型紫外分光光度計(jì),Perkin-Elmer683型紅外光譜儀,JNM-GX400型核磁共振儀,VGZ-AB-2F型質(zhì)譜儀,柱色譜和薄層色譜用硅膠、硅藻土(青島海洋化工廠分廠),柱色譜用聚酰胺和聚酰胺薄膜(浙江黃巖四青生化材料廠)。山海螺干切片購(gòu)于安國(guó)藥材市場(chǎng),經(jīng)中國(guó)中醫(yī)研究院謝宗萬(wàn)研究員鑒定為山海螺CodonopsislanceolataBenth.etHook。2c.n-fmoc-l-4-油醚、氯、醋酸乙酯萃取法取17kg山海螺粗粉,用95%工業(yè)乙醇溫浸,減壓回收乙醇,得浸膏?？偨嘤?kg硅藻土混合均勻,采用一般固液萃取方法,依次用石油醚、氯仿、醋酸乙酯萃取。分別回收萃取溶劑,得石油醚部分、氯仿部分、醋酸乙酯部分。石油醚部分120.5g,用200～300目硅膠常壓柱色譜,氯仿-醋酸乙酯梯度洗脫,重結(jié)晶精制得晶體Ⅰ、Ⅱ、Ⅲ;氯仿部分50g,經(jīng)多次硅膠柱色譜得晶體Ⅴ;醋酸乙酯部分20g,用80～100目聚酰胺柱色譜分離,水-丙酮梯度洗脫,聚酰胺薄膜TLC檢測(cè),合并相同部分,然后用硅膠柱色譜、重結(jié)晶方法純化精制得晶體Ⅳ、Ⅵ、Ⅶ。3化合物酮的測(cè)定化合物Ⅰ:無(wú)色針晶。mp213～214℃(未校正)。Liebermann-Burchard反應(yīng)呈陽(yáng)性,EI-MS(m/z):424(M+),409,300,285,257,218,205,204(基峰),133,95。1H-NMR(CDCl3)W:顯示8個(gè)甲基[0.831,0.916,0.938,0.956,1.067,1.082,1.088,1.139(each3H,s,8×CH3)],烯氫W5.5(1H,dd,H-15)。IRνmaxKBrcm-1:3402(KBr中的雜質(zhì)水峰),3047,2937,2857,1709,1464,1375。13C-NMR(CDCl3)W:38.346(C-1),34.129(C-2),217.525(C-3),47.570(C-4),55.779(C-5),19.958(C-6),35.100(C-7),38.874(C-8),48.698(C-9),37.738(C-10),17.438(C-11),35.765(C-12),37.689(C-13),157.609(C-14),117.187(C-15),36.666(C-16),37.534(C-17),48.795(C-18),40.633(C-19),28.788(C-20),33.072(C-21),33.347(C-22),26.102(C-23),21.472(C-24),14.796(C-25),29.912(C-26),25.555(C-27),29.847(C-28),33.565(C-29),21.336(C-30)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道的蒲公英萜酮一致?；衔铫?白色針晶。mp229～230℃。Liebermann-Burchard反應(yīng)呈陽(yáng)性。EI-MS(m/z):426(M+),411,302,287,218,204(基峰),189,135,95。1H-NMR(CDCl3)W:顯示8個(gè)甲基[0.803,0.822,0.908,0.908,0.928,0.951,1.001,1.09(each3H,s,8×CH3)],5.534(1H,dd,J=3.0,8.1Hz,H-15),3.192(1H,dd,J=3.1,10.1Hz,H-3)。13C-NMR(CDCl3)W:37.970(C-1),27.125(C-2),79.033(C-3),38.951(C-4),55.506(C-5),18.768(C-6),35.093(C-7),38.733(C-8),48.731(C-9),37.712(C-10),17.470(C-11),35.757(C-12),37.540(C-13),158.061(C-14),116.847(C-15),36.649(C-16),37.712(C-17),49.255(C-18),41.300(C-19),28.769(C-20),33.671(C-21),33.069(C-22),27.968(C-23),15.418(C-24),15.392(C-25),29.798(C-26),25.873(C-27),29.890(C-28),33.319(C-29),21.283(C-30)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道的蒲公英萜醇完全一致?；衔铫?白色羽狀晶體(醋酸乙酯)。mp177～178℃。Liebermann-Burchard反應(yīng)呈陽(yáng)性。EI-MS、1H-NMR、13C-NMR光譜數(shù)據(jù)與文獻(xiàn)報(bào)道的α-菠甾醇一致?；衔铫?白色棱狀結(jié)晶(甲醇-水)。mp133～136℃。水溶液呈酸性pH值為2。IR、UV、EI-MS、1H-NMR數(shù)據(jù)與文獻(xiàn)報(bào)道的順丁烯二酸一致。化合物Ⅴa、Ⅴb(晶體Ⅴ):白色晶體。mp33～34℃。。EI-MS(m/z):57(基峰),71,85,97,111,113,127,141,155,169,183,197,211,225,239,253,267,281,295,309,323,337,351,356,379,393。FAB-MS給出兩個(gè)分子離子峰410,408。1H-NMR(CDCl3)W:0.853(t,J=12.5Hz,末端甲基),1.254[多個(gè)H,S,(CH2)n],1.544[多個(gè)H,S,(CH2)n],2.292(t,R″-CH2-COOR′)。:1738,2954,2920,2846,1473,1464,1377,1173,729,719。410(M+)符合飽和脂肪酸酯通式(CnH2n+1COOCmH2m+1),推定410為二十六烷酸甲酯的分子離子峰。408(M+)符合烷烴通式(CnH2n+2),推定408為二十九烷酸甲酯的分子離子峰。綜上光譜初步確定晶體Ⅴ為二十六烷酸甲酯(Ⅴa)與正二十九烷(Ⅴb)的混合物?；衔铫鯽、Ⅵb(晶體Ⅵ):白色蠟狀晶體。mp56～57℃。。EI-MS(m/z):43,57(基峰),71,85,97,…,相隔14amu遞增,323,337,368,369,589,603,618,631(M-31),647(M-CH3),662,368峰明顯高于周圍各峰。368的強(qiáng)碎片峰為經(jīng)麥?zhǔn)现嘏?形成的穩(wěn)定碎片長(zhǎng)鏈脂肪酸部分,由此推斷脂肪酸為二十四烷酸。FAB-MS給出663.4(M+1),即M為662。高質(zhì)量端662符合飽和脂肪酸酯通式。此裂解方式與飽和脂肪酸酯裂解一致。:2956,2916,2848,1738,1473,1462,1377,1261,1097,1024,729,719。1H-NMR(CDCl3)W:0.854(t,J=11.5Hz,末端甲基),1.254[s,(CH2)n],1.624[s,(CH2)n],2.353(t,R′-CH2-COOR″)。綜上光譜可推出晶體Ⅵ為二十四碳酸二十一烷醇酯(Ⅵa)和四十四烷酸甲酯(Ⅵb)混合物。化合物Ⅶa、Ⅶb、Ⅶc(晶體Ⅶ):白色片狀(或粉末狀)晶體。mp269～271℃,Liebermann-urchard反應(yīng)呈陽(yáng)性,molish反應(yīng)界面有紫紅色環(huán)。:3388,2954,2871,1650,1464,1381,1367,1163,1076,1030,970,890,798。EI-MS:576,574,463,457,442,433,414(579-162),412(574-162),397,395,381,351,299,301,273,271,255,229,213,161,83,55。符合甾萜苷的質(zhì)譜裂解方式。HREI-MS:576.439909-C35H60O6,574.430606-C35H58O6。1H-NMR(CDCl3)W:5.138(m),5.0(m),3.6235(d,J=9.5Hz),4.206(d,J=8Hz),說明苷的構(gòu)型為β苷。13C-NMR(CDCl3)W:碳碳雙鍵:138.809,137.502,128.820,116.858;糖端基碳100.669;糖環(huán)碳(76.5,76.379,76.195,73.235,60.922);69.955為苷元C3信號(hào)。苷(晶體Ⅶ)用5%H2SO4稀甲醇水解,得苷元,苷元EI-MSm/z(%)給出兩個(gè)分子離子峰:414(100%),412(82%),399(40%),397(38%),369(30%),327(20%),300(25%),271(95%),273(85%),255(75%),246(40%),161(32%),107(42%),81(48%)。苷元:3369,2956,2870,1662,1462,1383,1369,1076,1030,970,930,845,796。苷元醋酐吡啶乙酰化,得苷元乙?；?