王 妍，梁旭博，趙珍珠

澤漆全草中二萜類成分研究

王 妍，梁旭博*，趙珍珠*

河南中醫(yī)藥大學(xué)藥學(xué)院，河南 鄭州 450046

研究澤漆全草的二萜類化學(xué)成分及其抗炎活性。用醋酸乙酯對(duì)澤漆的95%乙醇提取物進(jìn)行萃取得粗提物，然后采用大孔樹脂、硅膠、中壓制備色譜、Sephadex LH-20和高效液相色譜等多種色譜技術(shù)對(duì)醋酸乙酯部位分離純化，根據(jù)波譜數(shù)據(jù)及理化性質(zhì)鑒定化合物結(jié)構(gòu)。從澤漆的醋酸乙酯部位分離得到20個(gè)二萜，分別鑒定為euphoscopin A（1）、euphoscopin B（2）、euphoscopin C（3）、euphoscopin E（4）、euphorbiapene D（5）、euphornin A（6）、euphornin B（7）、euphornin（8）、helioscopianoid M（9）、euphoheliosnoid D（10）、2α-hydroxy helioscopinolide B（11）、helioscopinolide A（12）、helioscopinolide B（13）、helioscopinolide C（14）、helioscopinolide D（15）、helioscopinolide H（16）、helioscopinolide L（17）、-16β,17-dihydroxyatlsan-3-one（18）、20--acetylingenol（19）、altotibetol（20）。并且所有化合物均篩選了NO生成抑制活性?；衔?5～17、20為首次從澤漆中分離得到，所有化合物均報(bào)道于大戟屬的不同植物，說明大戟屬植物合成二萜的酶系具有高度同源性；活性結(jié)果顯示6、8～10和19顯示了微弱的抗炎活性。

澤漆；二萜；抗炎活性；euphornin A ；helioscopinolide D；altotibetol

大戟屬的植物種類繁多、生境復(fù)雜、變異性大，廣泛分布于世界各地[1]。二萜作為大戟屬植物的主要及特征性成分，因?yàn)槠浣Y(jié)構(gòu)多樣性和活性多樣性一直是研究熱點(diǎn)[2]。澤漆L.別名貓眼草、五朵云、五燈草、五風(fēng)草，隸屬于大戟科（Euphorbiaceae）大戟屬L.，生長(zhǎng)于亞洲、歐洲及北非等地區(qū)。澤漆性涼，味辛苦，全草入藥，有清熱、祛痰、利尿消腫及殺蟲之效[1]。臨床可用于治療結(jié)核性瘺管，細(xì)菌性痢疾，食道癌，治療無黃疸型傳染性肝炎和防治流行性腮腺炎等[2]?，F(xiàn)代研究表明，澤漆主要含二萜類、黃酮、三萜、甾醇、多酚類化合物[3-5]。二萜作為澤漆中主要的生物活性物質(zhì)，具有抗腫瘤、抗菌、清除自由基、抗炎等作用[6-10]。目前，從澤漆中分離的二萜已超過100個(gè)，涉及的骨架類型有13種[7-8,10-19]。近期，從該植物報(bào)道了3個(gè)5/10駢合雙環(huán)體系的新穎且具有抗炎活性的賈白欖烷二萜類[2]。為了更好地闡明澤漆的類藥效物質(zhì)基礎(chǔ)，發(fā)現(xiàn)更多的結(jié)構(gòu)新穎和活性顯著的二萜，本研究選擇澤漆作為研究對(duì)象，共從醋酸乙酯部位分離并鑒定了20個(gè)二萜，分別為euphoscopin A（1）、euphoscopin B（2）、euphoscopin C（3）、euphoscopin E（4）、euphorbiapene D（5）、euphornin A（6）、euphornin B（7）、euphornin（8）、helioscopianoid M（9）、euphoheliosnoid D（10）、2α-hydroxy helioscopinolide B（11）、helioscopinolide A（12）、helioscopinolide B（13）、helioscopinolide C（14）、helioscopinolide D（15）、helioscopinolide H（16）、helioscopinolide L（17）、-16β,17- dihydroxyatlsan-3-one（18）、20--acetylingenol（19）、altotibetol（20），化合物15、16、17、20為首次從澤漆中分離得到。并對(duì)這些化合物進(jìn)行了抗炎活性研究，部分化合物顯示了微弱的抗炎活性。

1 儀器與材料

Bruker Avance III 500 MHz型核磁共振儀（德國(guó)Bruker公司）；Agilent 1260 Series高效液相色譜儀（美國(guó)Agilent公司）；AB SCIEX Qtrap 5500三重四級(jí)桿質(zhì)譜儀（美國(guó)AB公司）；LC-52型高壓制備液相色譜儀（賽譜銳思科技有限公司）；FLEXA-HP中壓快速制備色譜儀HP-Q-P050（天津博納艾杰爾科技有限公司）；N-1300D-WB型旋轉(zhuǎn)蒸發(fā)儀（日本EYELA公司）；200-300目硅膠（青島海洋化工廠）；5 cm×10 cm薄層色譜硅膠板（青島海洋化工廠）；大孔樹脂Diaion HP-20（日本三菱化學(xué)）；RP18柱色譜填料（12 nm，粒徑50 μm）（日本YMC公司）；Sephadex LH-20（GE Healthcare公司）；ZORBAX RX-C8（250 mm×9.4 mm，5 μm）色譜柱（美國(guó)Agilent公司），YMC-PACK ODS-A（250 mm×20 mm，5 μm）色譜柱（日本YMC公司）；YMC-PACK ODS-A（250 mm×10 mm，5 μm）色譜柱（日本YMC公司）；色譜純甲醇（天津四友精細(xì)化學(xué)品有限公司）、色譜純乙腈（德國(guó)Merck公司）；分析純醋酸乙酯、石油醚、丙酮（天津富宇精細(xì)化學(xué)品有限公司）。小鼠單核巨噬細(xì)胞RAW264.7購自中科院上海細(xì)胞庫，DMEM培養(yǎng)基和胎牛血清購自BI公司；Griess Reagent、對(duì)照藥物-NMMA購自Sigma公司；脂多糖（lipopolysaccharide，LPS）購自索萊寶公司。

澤漆于2017年5月采自河南省開封市陳留鎮(zhèn)，經(jīng)河南中醫(yī)藥大學(xué)藥學(xué)院董誠明教授鑒定為大戟科植物澤漆L.，標(biāo)本（HFC 201705）收藏于河南中醫(yī)藥大學(xué)天然產(chǎn)物研究室。

2 提取與分離

澤漆干燥全草6.0 kg，95%乙醇冷浸提取4次，每次72 h。合并濾液，減壓濃縮后，加入蒸餾水混懸，用醋酸乙酯萃取4次，萃取液經(jīng)減壓濃縮得到醋酸乙酯部位浸膏420 g。醋酸乙酯部位經(jīng)硅膠柱色譜分離，石油醚-丙酮（20∶1、2∶1、1∶1）梯度洗脫，得到F（第1部分）、S（第2部分）、T（第3部分）3個(gè)洗脫部位。

S部位（130 g）經(jīng)大孔樹脂，洗脫劑為甲醇-水（20%、40%、60%、80%、100%）梯度洗脫，得11個(gè)組分Sa～Sk。組分Sd經(jīng)凝膠色譜柱Sephadex LH-20（甲醇）得到4個(gè)組分Sd1～Sd4，其中Sd3經(jīng)正相硅膠色譜柱以石油醚-丙酮（2∶1）等度洗脫得到Sd3a、Sd3b、Sd3c，其中Sd3a經(jīng)半制備HPLC（乙腈-水70∶30→100∶0，R= 30 min， 4 mL/min）分離純化得到化合物3（82.6 mg，R= 9.2 min）、8（1.6 mg，R= 17.6 min）；Sd4經(jīng)正相硅膠色譜柱以石油醚-丙酮（2∶1）等度洗脫得到4個(gè)組分Sd4a～Sd4d，Sd4b經(jīng)半制備HPLC（乙腈-水60∶40→80∶20，R= 50 min，5 mL/min）分離純化得到化合物4（3.1 mg，R= 28.7 min）、5（1.9 mg，R= 36.1 min）。組分Sf經(jīng)凝膠色譜柱Sephadex LH-20（甲醇）得到5個(gè)組分Sf1～Sf5，Sf3經(jīng)半制備HPLC（乙腈-水42∶58→100∶0，R= 40 min，4 mL/min）分離純化得到化合物13（7.7 mg，R= 17.3 min）、10（2.6 mg，R= 20.6 min）、9（2.1 mg，R= 25.4 min）；Sf4經(jīng)半制備HPLC（乙腈-水55∶45→80∶20，R= 30 min，5 mL/min）分離純化得到化合物6（2.3 mg，R= 9.8 min）、2（4.6 mg，R= 16.6 min）。

T部位（120 g）經(jīng)大孔樹脂，甲醇-水（10%、30%、60%、80%、100%）梯度洗脫，得11個(gè)組分T1～T10。組分T8、T9經(jīng)RP18色譜柱甲醇-水（50%、60%、70%、80%、90%、100%）得到4個(gè)組分T8a～T8d。T8a經(jīng)半制備HPLC（甲醇-水35∶65→100∶0，R= 60 min，10 mL/min）得6個(gè)組分T8a1～T8a6，T8a3經(jīng)半制備HPLC（乙腈-水30∶70→50∶50，R= 30 min，4 mL/min）分離純化得化合物15（2.8 mg，R= 25.1 min）；T8a4經(jīng)半制備HPLC（乙腈-水30∶70→50∶50，R= 30 min，4 mL/min）分離純化得化合物20（2.9 mg，R= 29.0 min）；T8a5經(jīng)半制備HPLC（乙腈∶水30∶70→55∶45，R= 30 min，4 mL/min）分離純化得化合物16（1.9 mg，R= 18.1 min）。T8b經(jīng)半制備HPLC（甲醇-水35∶65→100∶0，R= 60 min，10 mL/min）得4個(gè)組分T8b1～T8b4，T8b2經(jīng)半制備HPLC（乙腈-水28∶72→58∶42，R= 30 min，4 mL/min）分離純化得化合物17（4.1 mg，R= 30.5 min）；T8b3經(jīng)半制備HPLC（乙腈-水30∶70→75∶25，R= 40 min，4 mL/min）分離純化得到化合物19（3.9 mg，R= 35.2 min）。組分T10經(jīng)RP18色譜柱甲醇-水（40%、60%、80%、100%）梯度洗脫得17個(gè)組分T10a～T10r。T10f經(jīng)半制備HPLC（甲醇-水65∶35→80∶20，R= 30 min，10 mL/min）制備得T10f1～T10f8，T10f6經(jīng)半制備HPLC（乙腈-水35∶65→60∶40，R= 30 min，4 mL/min）分離純化得到化合物14（2.0 mg，R= 21.7 min）。T10h經(jīng)半制備HPLC（甲醇-水65∶35→90∶10，R= 40 min，10 mL/min）制備得到7個(gè)組分T10h1～T10h7，T10h3經(jīng)半制備HPLC（乙腈-水40∶60→70∶30，R= 35 min，4 mL/min）分離純化得化合物11（23.6 mg，R= 15.2 min）；T10h4經(jīng)半制備HPLC（乙腈-水47∶53→75∶25，R= 35 min，4 mL/min）分離純化得到化合物18（3.9 mg，R= 12.6 min）、12（7.3 mg，R= 17.9 min）、7（1.5 mg，R= 24.2 min）；T10h5經(jīng)半制備HPLC（乙腈-水50∶50→75∶25，R= 35 min，4 mL/min）分離純化得到化合物1（2.1 mg，R= 32.8 min）。

3 結(jié)構(gòu)鑒定

化合物1：無色油狀物，ESI-MS/: 541 [M＋H]+，分子式為C31H40O8。1H-NMR (500 MHz, CDCl3): 7.98 (2H, d,= 7.8 Hz, H-3?, 7?), 7.56 (1H, t,= 7.4 Hz, H-5?), 7.44 (2H, t,= 7.7 Hz, H-4?, 6?), 5.91 (1H, s, H-14), 5.64 (1H, d,= 8.9 Hz, H-5), 5.30 (1H, d,= 16.0 Hz, H-11), 5.22 (1H, dd,= 7.2, 3.6 Hz, H-3), 5.11 (1H, dd,= 15.9, 8.9 Hz, H-12), 4.42 (1H, d,= 4.2 Hz, H-7), 3.28 (1H, t,= 8.2 Hz, H-4), 2.99 (1H, dd,= 15.0, 8.5 Hz, H-1), 2.96 (1H, dd,= 15.7, 10.1 Hz, H-8), 2.65 (1H, dd,= 15.5, 4.4 Hz, H-8), 2.39 (1H, m, H-2), 2.25 (1H, m, H-13), 2.18 (3H, s, 15-OCOCH3), 2.15 (3H, s, 14-OCOCH3), 1.82 (3H, s, 17-CH3), 1.43 (1H, dd,= 15.1, 9.1 Hz, H-1), 1.21 (3H, s, 19-CH3), 1.09 (3H, d,= 7.1 Hz, 20-CH3), 1.07 (3H, s, 18-CH3), 0.90 (3H, d,= 7.1 Hz, 16-CH3)；13C-NMR (125 MHz, CDCl3): 209.6 (C-9), 170.2 (14, 15-OCOCH3), 166.4 (C-1?), 140.2 (C-6), 134.0 (C-11), 133.4 (C-5?), 133.1 (C-12), 130.3 (C-2?), 129.5 (C-3?, 7?), 128.7 (C-4?, 6?), 120.7 (C-5), 92.2 (C-15), 83.7 (C-3), 75.6 (C-14), 71.8 (C-7), 49.2 (C-10), 45.5 (C-4), 44.6 (C-1), 42.9 (C-8), 38.1 (C-2), 37.2 (C-13), 25.5 (C-18), 25.1 (C-19), 22.9 (C-20), 22.2 (15-OCOCH3), 21.2 (14-OCOCH3), 18.9 (C-16), 18.7 (C-17)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[14]，故鑒定化合物1為euphoscopin A。

化合物3：無色油狀物，ESI-MS/: 645 [M＋H]+，分子式為C38H44O9。1H-NMR (500 MHz, CDCl3): 7.85 (2H, d,= 7.4 Hz, H-3?, 7?), 7.56 (2H, d,= 7.5 Hz, H-3??, 7??), 7.46 (1H, t,= 7.4 Hz, H-5?), 7.30 (3H, dd,= 12.4, 7.6 Hz, H-4?, 6?, 5??), 6.97 (2H, t,= 7.7 Hz, H-4??, 6??), 5.94 (1H, s, H-14), 5.86 (1H, d,= 8.7 Hz, H-5), 5.70 (1H, dd,= 11.4, 4.2 Hz, H-7), 5.39 (1H, d,= 16.0 Hz, H-11), 5.21 (1H, dd,= 16.0, 8.9 Hz, H-12), 5.13 (1H, dd,= 6.5, 2.0 Hz, H-3), 3.32 (1H, m, H-8), 3.29 (1H, m, H-4), 3.00 (1H, dd,= 15.5, 8.4 Hz, H-1), 2.85 (1H, dd,= 15.8, 4.2 Hz, H-8), 2.46 (1H, m, H-2), 2.20 (3H, s, 15-OCOCH3), 2.17 (3H, s, 14-OCOCH3), 2.13 (1H, m, H-13), 1.95 (3H, s, 17-CH3), 1.49 (1H, dd,= 15.5, 7.8 Hz, H-1), 1.31 (3H, s, 19-CH3), 1.14 (3H, s, 18-CH3), 1.09 (3H, d,= 7.2 Hz, 20-CH3), 0.93 (3H, d,= 7.1 Hz, 16-CH3)；13C-NMR (125 MHz, CDCl3): 207.6 (C-9), 170.1 (14-OCOCH3), 170.1 (15-OCOCH3), 165.9 (C-1?), 165.5 (C-1??), 135.5 (C-6), 133.7 (C-11), 133.7 (C-5?), 132.7 (C-5??), 132.5 (C-12), 130.5 (C-2?), 129.9 (C-2??), 129.4 (C-3?, 7?), 129.3 (C-3??, 7??), 128.3 (C-4?, 6?), 128.0 (C-4??, 6??), 122.8 (C-5), 92.5 (C-15), 84.3 (C-3), 75.7 (C-14), 74.2 (C-7), 49.2 (C-10), 44.1 (C-4), 43.7 (C-1), 42.9 (C-8), 38.0 (C-2), 37.9 (C-13), 25.5 (C-18), 25.0 (C-19), 22.9 (C-20), 22.1 (15-OCOCH3), 21.1 (14-OCOCH3), 19.3 (C-16), 18.9 (C-17)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[15]，故鑒定化合物3為euphoscopin C。

化合物4：無色油狀物，ESI-MS/: 497 [M＋H]+，分子式為C29H36O7。1H-NMR (500 MHz, CDCl3): 8.03 (2H, dd,= 8.3, 1.3 Hz, H-3?, 7?), 7.58 (1H, t,= 7.4 Hz, H-5?), 7.46 (1H, t,= 7.7 Hz, H-4?, 6?), 5.81(1H, d,= 10.4 Hz, H-5), 5.49 (1H, d,= 15.6 Hz, H-11), 5.18 (1H, dd,= 6.6, 2.9 Hz, H-3), 5.08 (1H, dd,= 15.6, 9.0 Hz, H-12), 4.36 (1H, m, H-7), 3.40 (1H, m, H-13), 3.12 (1H, dd,= 9.6, 6.6 Hz, H-4), 2.88 (1H, dd,= 14.4, 10.2 Hz, H-8), 2.71 (1H, m, H-8), 2.67 (1H, m, H-1), 2.34 (1H, m, H-2), 2.30 (3H, s, 15-OCOCH3), 2.24 (1H, m, H-1), 1.55 (3H, s, 17-CH3), 1.22 (3H, d,= 7.1 Hz, 16-CH3), 1.17 (3H, s, 19-CH3), 1.12 (3H, d,= 6.6 Hz, 20-CH3), 1.11 (3H, s, 18-CH3)；13C-NMR (125 MHz, CDCl3): 211.8 (C-14), 209.3 (C-9), 171.0 (15-OCOCH3), 166.2 (C-1?), 143.5 (C-6), 136.4 (C-11), 133.4 (C-5?), 131.2 (C-12), 130.3 (C-2?), 129.6 (C-3?, 7?), 128.7 (C-4?, 6?), 117.8 (C-5), 96.1 (C-15), 84.0 (C-3), 72.0 (C-7), 51.5 (C-4), 49.6 (C-10), 45.8 (C-13), 45.1 (C-8), 42.5 (C-1), 39.0 (C-2), 25.8 (C-19), 22.0 (C-18), 21.9 (15-OCOCH3), 18.8 (C-16), 18.7 (C-20), 18.3 (C-17)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[16]，故鑒定化合物4為euphoscopin E。

化合物5：白色無定形粉末，ESI-MS/: 623 [M＋Na]+，分子式為C36H40O8。1H-NMR (500 MHz, CDCl3): 7.85 (2H, d,= 7.3 Hz, H-3??, 7??), 7.61 (2H, d,= 7.4 Hz, H-3?, 7?), 7.48 (1H, t,= 7.4 Hz, H-5??), 7.29 (3H, t,= 7.7 Hz, H-4??, 6??, 5?), 6.94 (2H, t,= 7.8 Hz, H-4?, 6?), 5.88 (1H, d,= 9.2 Hz, H-5), 5.58 (1H, d,= 15.5 Hz, H-11), 5.55 (1H, dd,= 11.3, 4.3 Hz, H-7), 5.11 (1H, dd,= 6.1, 1.3 Hz, H-3), 5.07 (1H, dd,= 15.6, 9.5 Hz, H-12), 3.47 (1H, dd,= 9.4, 6.7 Hz, H-4), 3.24 (1H, dd,= 14.8, 11.4 Hz, H-8), 3.14 (1H, dd,= 9.0, 6.1 Hz, H-13), 2.89 (1H, dd,= 14.8, 4.5 Hz, H-8), 2.65 (1H, dd,= 14.0, 7.2 Hz, H-1), 2.33 (3H, s, 15-OCOCH3), 2.24 (1H, m, H-2), 2.22 (1H, m, H-1), 1.77 (3H, s, 17-CH3), 1.27 (3H, s, 19-CH3), 1.22 (3H, d,= 6.9 Hz, 16-CH3), 1.15 (6H, t,= 3.2 Hz, 18, 20-CH3)；13C-NMR (125 MHz, CDCl3): 212.0 (C-14), 206.7 (C-9), 171.0 (15-OCOCH3), 165.4 (C-1?, 1??), 139.8 (C-6), 136.5 (C-11), 132.9 (C-5?), 132.7 (C-5??), 132.6 (C-12), 130.4 (C-2?), 129.9 (C-2??), 129.5 (C-3??, 7??), 129.3 (C-3?, 7?), 128.4 (C-4??, 6??), 128.1 (C-4?, 6?), 119.5 (C-5), 96.3 (C-15), 84.4 (C-3), 74.1 (C-7), 51.3 (C-13), 49.5 (C-10), 45.8 (C-4), 43.8 (C-8), 43.1 (C-1), 39.6 (C-2), 25.3 (C-19), 22.5 (C-20), 22.0 (15-OCOCH3), 19.2 (C-17, 18), 19.0 (C-16)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[17]，故鑒定化合物5為euphorbiapene D。

化合物6：無色油狀物，ESI-MS/: 543 [M＋H]+，分子式為C31H42O8。1H-NMR (500 MHz, CDCl3): 8.06 (2H, dd,= 8.3, 1.3 Hz, H-3?, 7?), 7.53 (1H, t,= 7.4 Hz, H-5?), 7.43 (2H, t,= 7.7 Hz, H-4?, 6?), 5.82 (1H, d,= 10.5 Hz, H-5), 5.62 (1H, dd,= 15.6, 9.5 Hz, H-12), 5.40 (1H, t,= 4.3 Hz, H-3), 5.06 (1H, d,= 15.6 Hz, H-11), 4.93 (1H, d,= 2.8 Hz, H-14), 4.36 (1H, dd,= 5.2, 1.7 Hz, H-7), 4.09 (1H, t,= 4.5 Hz, H-9), 2.97 (1H, dd,= 10.7, 5.2 Hz, H-4), 2.60 (1H, m, H-13), 2.22 (3H, brs, 14-OCOCH3), 2.17 (1H, m, H-2), 2.05 (3H, s, 9-OCOCH3), 2.00 (1H, m, H-1), 1.98 (1H, m, H-8), 1.77 (1H, m, H-1), 1.74 (1H, m, H-8), 1.68 (3H, d,= 1.0 Hz, 17-CH3), 1.03 (3H, s, 19-CH3), 0.97 (3H, d,= 6.7 Hz, 16-CH3), 0.95 (3H, d,= 7.0 Hz, 20-CH3), 0.94 (3H, s, 18-CH3)；13C-NMR (125 MHz, CDCl3): 172.1 (14-OCOCH3), 171.5 (9-OCOCH3), 166.5 (C-1?), 138.6 (C-11), 137.8 (C-6), 132.9 (C-5?), 130.4 (C-2?), 129.9 (C-3?, 7?), 129.0 (C-12), 128.5 (C-4?, 6?), 119.1 (C-5), 83.9 (C-15), 81.2 (C-3), 80.9 (C-14), 75.5 (C-7), 71.9 (C-9), 47.8 (C-4), 45.8 (C-1), 39.6 (C-10), 39.4 (C-13), 37.0 (C-2), 35.2 (C-8), 23.2 (14-OCOCH3), 21.5 (C-19), 21.2 (9-OCOCH3), 20.4 (C-20), 19.6(C-18), 16.6 (C-17), 13.9 (C-16)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[16]，故鑒定化合物6為euphornin A。

化合物7：無色油狀物，ESI-MS/: 543 [M＋H]+，分子式為C31H42O8。1H-NMR (500 MHz, CDCl3): 8.07 (2H, d,= 7.7 Hz, H-3?, 7?), 7.54 (1H, t,= 7.0 Hz, H-5?), 7.44 (2H, t,= 7.6 Hz, H-4?, 6?), 5.59 (1H, dd,= 15.5, 9.3 Hz, H-12), 5.49 (1H, d,= 10.8 Hz, H-5), 5.42 (1H, t,= 3.7 Hz, H-3), 5.08 (2H, m, H-7, 11), 4.93 (1H, d,= 2.1 Hz, H-14), 3.33 (1H, t,= 4.8 Hz, H-9), 2.87 (1H, dd,= 10.3, 4.6 Hz, H-4), 2.54 (1H, m, H-13), 2.23 (3H, s, 14-OCOCH3), 2.14 (1H, m, H-2), 2.03 (2H, m, H-1, 8), 1.96 (1H, m, H-8), 1.78 (1H, m, H-1), 1.73 (3H, s, 17-CH3), 1.25 (3H, s, 7-OCOCH3), 1.11 (3H, s, 18-CH3), 0.95 (6H, d,= 6.6 Hz, 16, 20-CH3), 0.85 (3H, s, 19-CH3)；13C-NMR (125 MHz, CDCl3): 173.1 (14-OCOCH3), 171.2 (7-OCOCH3), 165.4 (C-1?), 140.3 (C-11), 134.5 (C-6), 132.9 (C-5?), 130.2 (C-2?), 129.7 (C-3?, 7?), 128.4 (C-12), 127.7 (C-4?, 6?), 119.2 (C-5), 84.0 (C-15), 80.9 (C-3), 80.7 (C-14), 73.6 (C-9), 73.3 (C-7), 47.8 (C-4), 45.9 (C-1), 40.2 (C-10), 39.5 (C-13), 36.6 (C-2), 34.8 (C-8), 22.8 (C-18), 21.0 (14-OCOCH3), 20.4 (7-OCOCH3), 20.0 (C-19), 18.6 (C-20), 16.3 (C-17), 13.4 (C-16)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[18]，故鑒定化合物7為euphornin B。

化合物8：白色針晶（甲醇），ESI-MS/: 585 [M＋H]+，分子式為C33H44O9。1H-NMR (500 MHz, CDCl3): 8.08 (2H, d,= 7.1 Hz, H-3?, 7?), 7.52 (1H, dd,= 10.5, 4.2 Hz, H-5?), 7.44 (2H, t,= 7.5 Hz, H-4?, 6?), 5.70 (1H, d,= 10.3 Hz, H-5), 5.62 (1H, dd,= 15.5, 9.3 Hz, H-12), 5.42 (1H, t,= 4.1 Hz, H-3), 5.05 (1H, d,= 15.6 Hz, H-11), 4.93 (1H, t,= 4.0 Hz, H-14), 4.76 (1H, t,= 3.6 Hz, H-9), 2.88 (1H, dd,= 10.3, 4.8 Hz, H-4), 2.55 (1H, m, H-13), 2.22 (3H, s, 14-OCOCH3), 2.13 (1H, m, H-2), 2.05 (1H, m, H-1), 1.89～2.00 (2H, m, H-2), 1.95 (3H, s, 9-OCOCH3), 1.77 (1H, m, H-1), 1.72 (3H, brs, 17-CH3), 1.17 (3H, brs, 7-OCOCH3), 0.95 (9H, t,= 5.5 Hz, 16, 18, 20-CH3), 0.88 (3H, s, 19-CH3)；13C-NMR (125 MHz, CDCl3): 171.4 (14-OCOCH3), 169.8 (7-OCOCH3), 169.2 (9-OCOCH3), 165.8 (C-1?), 138.4 (C-11), 133.9 (C-6), 133.0 (C-5?), 130.3 (C-2?), 129.9 (C-3?, 7?), 128.7 (C-12), 128.6 (C-4?, 6?), 120.2 (C-5), 83.9 (C-15), 81.1 (C-3), 80.8 (C-14), 73.7 (C-9), 73.0 (C-7), 48.0 (C-4), 46.3 (C-1), 39.8 (C-10), 39.6 (C-13), 36.8 (C-2), 32.5 (C-8), 22.7 (C-18), 21.2 (9-OCOCH3), 21.1 (14-OCOCH3), 20.3 (C-19), 20.0 (7-OCOCH3), 19.6 (C-20), 16.3 (C-17), 13.6 (C-16)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[16]，故鑒定化合物8為euphornin。

化合物9：無色油狀物，ESI-MS/: 519 [M＋Na]+，分子式為C29H36O7。1H-NMR (500 MHz, CDCl3): 7.94 (1H, d,= 7.1 Hz, H-3?, 7?), 7.60 (1H, t,= 7.4 Hz, H-5?), 7.49 (1H, t,= 7.7 Hz, H-4?, 6?), 5.66 (1H, d,= 15.8 Hz, H-11), 5.44 (2H, dd,= 10.7, 4.3 Hz, H-3, 5), 5.27 (1H, dd,= 15.9, 9.3 Hz, H-12), 5.11 (1H, brs, H-17), 4.98 (1H, brs, H-17), 4.05 (1H, m, H-13), 3.49 (1H, dd,= 10.5, 4.2 Hz, H-4), 2.59 (1H, m, H-7), 2.54 (1H, m, H-8), 2.42 (2H, m, H-2, 8), 2.26 (1H, dd,= 14.8, 8.6 Hz, H-1), 2.21 (1H, m, H-7), 1.86 (3H, s, 5-OCOCH3), 1.78 (1H, dd,= 14.7, 3.3 Hz, H-1), 1.25 (3H, s, 18-CH3), 1.19 (3H, s, 19-CH3), 1.17 (3H, d,= 6.7 Hz, 20-CH3), 1.14 (3H, d,= 7.3 Hz, 16-CH3)；13C-NMR (125 MHz, CDCl3): 211.9 (C-14), 211.2 (C-9), 170.2 (5-OCOCH3), 165.8 (C-1?), 145.1 (C-6), 137.4 (C-11), 133.5 (C-5?), 131.0 (C-12), 130.0 (C-2?), 129.6 (C-3?, 7?), 128.9 (C-4?, 6?), 115.4 (C-17), 88.6 (C-15), 84.8 (C-3), 70.0 (C-5), 50.7 (C-1), 50.0 (C-10), 49.2 (C-4), 43.7 (C-13), 38.0 (C-2), 35.4 (C-8), 30.0 (C-7), 24.3 (C-18), 24.0 (C-19), 21.1 (5-OCOCH3), 18.9 (C-16), 17.6 (C-20)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[19]，故鑒定化合物9為helioscopianoid M。

由圖2可以看出，隨著升溫速率的增加，DTG曲線峰值向較高溫度區(qū)域移動(dòng)，并且揮發(fā)分析出的溫度范圍(失重第二階段)也呈現(xiàn)增加趨勢(shì)，這可以促進(jìn)污泥內(nèi)部有機(jī)物質(zhì)的熱分解反應(yīng)的進(jìn)行。同時(shí)，升溫速率的增加可以明顯提高污泥熱解的最大失重速率。這主要是由于升溫速率的增加縮短了熱解反應(yīng)時(shí)間，從而影響了污泥熱解過程中化學(xué)轉(zhuǎn)化反應(yīng)的發(fā)生。同時(shí)，在較高的升溫速率下，由于傳熱與傳質(zhì)擴(kuò)散的影響，污泥樣品與熱重儀反應(yīng)室中的溫差也會(huì)變大，樣品內(nèi)部顆粒的溫度和外部顆粒的溫差增大，揮發(fā)分的析出釋放峰滯后，從而使污泥的轉(zhuǎn)化率降低。因此，一定程度上可以通過提高污泥熱解的升溫速率達(dá)到提高污泥整體處理效率的目的。

化合物10：無色油狀物，ESI-MS/: 513 [M＋H]+，分子式為C29H36O8。1H-NMR (500 MHz, CDCl3): 8.04 (2H, d,= 8.0 Hz, H-3?, 7?), 7.56 (1H, t,= 7.1 Hz, H-5?), 7.45 (1H, t,= 7.7 Hz, H-4?, 6?), 6.37 (1H, d,= 8.7 Hz, H-12), 5.93 (1H, d,= 11.0 Hz, H-5), 4.99 (1H, t,= 6.3 Hz, H-3), 4.21 (1H, d,= 8.7 Hz, H-11), 4.08 (1H, m, H-7), 3.66 (1H, d,= 17.1 Hz, H-8), 3.28 (1H, dd,= 10.9, 7.5 Hz, H-1), 2.64 (1H, dd,= 14.6, 9.0 Hz, H-4), 2.46 (2H, m, H-2, 8), 2.19 (3H, s, 15-OCOCH3), 1.83 (3H, s, 20-CH3), 1.49 (3H, brs, 17-CH3), 1.28 (3H, s, 18-CH3), 1.16 (3H, d,= 6.8 Hz, 16-CH3), 0.90 (3H, s, 19-CH3)；13C-NMR (125 MHz, CDCl3): 219.5 (C-9), 200.7 (C-14), 170.8 (15-OCOCH3), 166.4 (C-1?), 142.9 (C-6), 138.9 (C-13), 135.1 (C-12), 133.1 (C-5?), 130.6 (C-2?), 129.7 (C-3?, 7?), 128.5 (C-4?, 6?), 118.0 (C-5), 93.3 (C-15), 83.1 (C-3), 74.0 (C-7), 52.8 (C-10), 48.1 (C-1), 41.8 (C-4), 40.4 (C-8), 37.5 (C-2), 22.5 (C-19), 21.6 (15-OCOCH3), 20.4 (C-18), 17.2 (C-17), 15.7 (C-16), 12.2 (C-20)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[20]，故鑒定化合物10為euphoheliosnoid D。

化合物11：黃色針晶（甲醇），ESI-MS/: 333 [M＋H]+，分子式為C20H28O4。1H-NMR (500 MHz, CDCl3): 6.30 (1H, s, H-14), 4.94 (1H, dd,= 13.1, 5.7 Hz, H-12), 4.20 (1H, d,= 3.0 Hz, H-2), 3.21 (1H, d,= 2.1 Hz, H-3), 2.63 (1H, dd,= 13.4, 6.2 Hz, H-11), 2.53 (1H, brd,= 12.7 Hz, H-7), 2.40 (1H, dd,= 14.3, 2.9 Hz, H-1), 2.21 (1H, td,= 13.0, 4.3 Hz, H-7), 2.16 (1H, d,= 8.7 Hz, H-9), 1.88 (1H, m, H-6), 1.82 (3H, s, 20-CH3), 1.55 (1H, m, H-6), 1.53 (1H, m, H-11), 1.40 (1H, dd,= 14.3, 2.5 Hz, H-1), 1.22 (1H, dd,= 12.3, 1.5 Hz, H-5), 1.17 (3H, s, 19-CH3), 1.03 (6H, d,= 4.6 Hz, 17-CH3, 18-CH3)；13C-NMR (125 MHz, CDCl3): 175.9 (C-16), 156.7 (C-13), 151.8 (C-8), 116.5 (C-15), 114.6 (C-14), 78.1 (C-3), 76.4 (C-12), 70.7 (C-2), 54.4 (C-5), 52.5 (C-9), 43.3 (C-1), 40.7 (C-10), 38.7 (C-4), 37.1 (C-7), 30.5 (C-17), 27.7 (C-11), 23.4 (C-6), 18.5 (C-19), 17.3 (C-18), 8.4 (C-20)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[20]，故鑒定化合物11為2α-hydroxy helioscopinolide B。

化合物12：無色油狀物，ESI-MS/: 317 [M＋H]+，分子式為C20H28O3。1H-NMR (500 MHz, CDCl3): 6.28 (1H, s, H-14), 4.86 (1H, dd,= 13.3, 5.9 Hz, H-12), 3.28 (1H, dd,= 11.8, 4.2 Hz, H-3), 2.55 (1H, dd,= 13.5, 5.8 Hz, H-11), 2.51 (1H, m, H-7), 2.21 (1H, dd,= 13.1, 5.0 Hz, H-7), 2.16 (1H, d,= 8.2 Hz, H-9), 1.96 (1H, dt,= 12.9, 3.1 Hz, H-1), 1.83 (3H, s, 20-CH3), 1.76 (1H, m, H-2), 1.62 (1H, ddd,= 15.9, 13.4, 3.2 Hz, H-2), 1.52 (1H, m, H-11), 1.44 (1H, m, H-6), 1.25 (1H, td,= 13.2, 3.1 Hz, H-1), 1.15 (1H, dd,= 12.4, 1.8 Hz, H-5), 1.03 (3H, s, 17-CH3), 0.93 (3H, s, 19-CH3), 0.82 (3H, s, 18-CH3)；13C-NMR (125 MHz, CDCl3): 175.4 (C-16), 156.2 (C-13), 151.5 (C-8), 116.7 (C-15), 114.4 (C-14), 78.7 (C-3), 76.0 (C-12), 54.5 (C-5), 51.7 (C-9), 41.4 (C-10), 39.2 (C-4), 37.6 (C-1), 37.1 (C-7), 28.8 (C-17), 27.8 (C-2), 27.7 (C-11), 23.6 (C-6), 16.9 (C-19), 15.7 (C-18), 8.4 (C-20)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[21]，故鑒定化合物12為helioscopinolide A。

化合物13：黃色針晶（甲醇），ESI-MS/: 317 [M＋H]+，分子式為C20H28O3。1H-NMR (500 MHz, CDCl3): 6.27 (1H, s, H-14), 4.88 (1H, dd,= 13.1, 5.8 Hz, H-12), 3.49 (1H, t,= 2.7 Hz, H-3), 2.58 (1H, dd,= 13.6, 6.2 Hz, H-11), 2.51 (1H, m, H-7), 2.30 (1H, d,= 8.6 Hz, H-9), 2.24 (1H, m, H-7), 1.97 (1H, m, H-1), 1.83 (3H, s, 20-CH3), 1.70～1.77 (2H, m, H-2), 1.71 (1H, m, H-6), 1.66 (2H, m, H-1, 5), 1.51 (1H, m, H-11), 1.44 (1H, dd,= 12.9, 4.1 Hz, H-6), 1.00 (3H, s, 17-CH3), 0.95 (3H, s, 19-CH3), 0.88 (3H, s, 18-CH3)；13C-NMR (125 MHz, CDCl3): 175.6 (C-16), 156.3 (C-13), 152.2 (C-8), 116.5 (C-15), 114.2 (C-14), 76.2 (C-12), 75.8 (C-3), 51.7 (C-9), 48.4 (C-5), 41.4 (C-10), 37.9 (C-4), 37.2 (C-7), 32.2 (C-1), 28.9 (C-17), 27.6 (C-11), 25.8 (C-2), 23.5 (C-6), 22.4 (C-18), 16.9 (C-19), 8.4 (C-20)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[22]，故鑒定化合物13為helioscopinolide B。

化合物14：白色針晶（甲醇），ESI-MS/: 331 [M＋H]+，分子式為C20H26O4。1H-NMR (500 MHz, CDCl3): 6.35 (1H, s, H-14), 4.85 (1H, dd,= 13.2, 5.9 Hz, H-12), 3.97 (1H, d,= 4.7 Hz, H-3), 3.42 (1H, d,= 4.9 Hz, 3-OH), 2.74 (1H, d,= 12.4 Hz, H-1), 2.60 (1H, brd,= 13.7 Hz, H-7), 2.50 (1H, d,= 8.6 Hz, H-9), 2.41 (1H, dd,= 14.5, 6.9 Hz, H-11), 2.38 (1H, d,= 13.3 Hz, H-1), 2.30 (1H, td,= 13.3, 4.9 Hz, H-7), 1.98 (1H, dt,= 13.3, 2.5 Hz, H-6), 1.85 (3H, s, 20-CH3), 1.82 (1H, brd,= 12.6 Hz, H-5), 1.63 (1H, m, H-11), 1.52 (1H, ddd,= 17.1, 13.4, 4.2 Hz, H-6), 1.23 (3H, s, 17-CH3), 0.92 (3H, s, 19-CH3), 0.71 (3H, s, 18-CH3)；13C-NMR (125 MHz, CDCl3): 209.5 (C-2), 175.0 (C-16), 155.1 (C-13), 149.3 (C-8), 117.8 (C-15), 115.5 (C-14), 82.6 (C-3), 75.5 (C-12), 53.7 (C-5), 51.5 (C-9), 51.4 (C-1), 47.1 (C-10), 45.3 (C-4), 36.5 (C-7), 29.7 (C-17), 27.8 (C-11), 23.2 (C-6), 17.5 (C-19), 16.6 (C-18), 8.5 (C-20)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[21]，故鑒定化合物14為helioscopinolide C。

化合物15：無色油狀物，ESI-MS/: 331 [M＋H]+，分子式為C20H26O4。1H-NMR (500 MHz, CDCl3): 6.56 (1H, s, H-14), 4.79 (1H, d,= 12.0 Hz, H-12), 3.02 (1H, m, H-11), 2.98 (1H, m, H-7), 2.6-2.7 (2H, m, H-2), 2.57 (2H, m, H-5, 7), 2.39 (1H, m, H-1), 1.94 (1H, m, H-1), 1.90 (3H, s, 20-CH3), 1.72 (1H, m, H-6), 1.54 (1H, m, H-6), 1.39 (1H, m, H-11), 1.37 (3H, s, 17-CH3), 1.31 (3H, s, 19-CH3), 1.25 (3H, s, 18-CH3)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[21]，故鑒定化合物15為helioscopinolide D。

化合物16：白色無定形粉末，ESI-MS/: 333 [M＋H]+，分子式為C20H28O4。1H-NMR (500 MHz, CDCl3): 6.37 (1H, s, H-14), 4.88 (1H, dd,= 12.7, 6.0 Hz, H-12), 3.31 (1H, dd,= 12.0, 3.5 Hz, H-3), 3.10 (1H, dd,= 13.4, 6.3 Hz, H-11), 2.70 (1H, m, H-7), 2.31 (1H, m, H-7), 1.89 (2H, m, H-2, 5), 1.86 (3H, s, 20-CH3), 1.78 (2H, m, H-2, 6), 1.65 (1H, m, H-1), 1.62 (1H, m, H-1), 1.42 (1H, m, H-6), 1.33 (1H, m, H-11), 1.06 (3H, s, 17-CH3), 0.99 (3H, s, 19-CH3), 0.85 (3H, s, 18-CH3)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[22]，故鑒定化合物16為helioscopinolide H。

化合物17：白色無定形粉末，ESI-MS/: 333 [M＋H]+，分子式為C19H24O5。1H-NMR (500 MHz, CDCl3): 6.38 (1H, s, H-14), 5.69 (1H, s, H-1), 4.94 (1H, dd,= 13.6, 5.8 Hz, H-12), 2.87 (1H, d,= 7.9 Hz, H-9), 2.54 (2H, m, H-7, 11), 2.40 (1H, brd,= 12.9 Hz, H-5), 2.25 (1H, td,= 13.4, 4.7 Hz, H-7), 1.85 (3H, s, 20-CH3), 1.81 (1H, m, H-6), 1.53 (1H, m, H-11), 1.50 (1H, m, H-6), 1.36 (3H, s, 17-CH3), 1.24 (3H, s, 18-CH3), 1.05 (3H, s, 19-CH3)；13C-NMR (125 MHz, CDCl3): 177.7 (C-3), 176.0 (C-16), 156.2 (C-13), 150.7 (C-8), 117.3 (C-15), 115.4 (C-14), 98.9 (C-1), 76.1 (C-12), 43.5 (C-9), 43.0 (C-10), 42.9 (C-5), 40.5 (C-4), 36.1 (C-7), 29.7 (C-17), 27.3 (C-11), 24.3 (C-18), 24.2 (C-6), 14.6 (C-19), 8.4 (C-20)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[22]，故鑒定化合物17為helioscopinolide L。

化合物18：黃色針晶（甲醇），ESI-MS/: 321 [M＋H]+，分子式為C20H32O3。1H-NMR (500 MHz, CDCl3): 3.58 (1H, d,= 10.9 Hz, H-17), 3.44 (1H, d,= 10.9 Hz, H-17), 2.58 (1H, ddd,= 16.0, 12.4, 6.9 Hz, H-2), 2.34 (1H, ddd,= 16.0, 6.0, 3.2 Hz, H-2), 2.02 (1H, m, H-11), 1.87 (1H, m, H-14), 1.84 (2H, m, H-1, 12), 1.62 (1H, m, H-13), 1.50 (1H, m, H-13), 1.46 (2H, m, H-6), 1.43 (1H, m, H-7), 1.37 (1H, m, H-1), 1.35 (1H, m, H-9), 1.31 (1H, m, H-5), 1.23 (2H, m, H-11, 15), 1.16 (1H, m, H-7), 1.11 (3H, s, 20-CH3), 1.10 (1H, m, H-15), 1.08 (3H, s, 18-CH3), 1.04 (3H, s, 19-CH3), 0.82 (1H, m, H-14)；13C-NMR (125 MHz, CDCl3): 217.6 (C-3), 74.2 (C-16), 69.1 (C-17), 55.8 (C-5), 52.6 (C-15), 51.0 (C-9), 47.8 (C-4), 38.9 (C-7), 38.1 (C-1), 37.3 (C-10), 34.2 (C-2), 33.0 (C-8), 32.3 (C-12), 27.3 (C-14), 26.3 (C-18), 23.3 (C-13), 23.1 (C-11), 21.8 (C-19), 19.8 (C-6), 13.6 (C-20)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[23]，故鑒定化合物18為-16β,17-dihydroxyatlsan-3-one。

化合物19：無色油狀物，ESI-MS/: 391 [M＋H]+，分子式為C22H30O6。1H-NMR (500 MHz, CDCl3): 6.10 (1H, d,= 4.1 Hz, H-7), 5.94 (1H, s, H-1), 4.71 (1H, d,= 12.6 Hz, H-20), 4.51 (1H, d,= 12.6 Hz, H-20), 4.43 (1H, s, H-3), 4.09 (1H, d,= 11.6 Hz, H-8), 3.67 (1H, s, H-5), 2.31 (1H, m, H-11), 2.26 (1H, m, H-12), 2.05 (3H, s, 20-OCOCH3), 1.85 (3H, s, 19-CH3), 1.76 (1H, m, H-12), 1.11 (3H, s, 17-CH3), 1.06 (3H, s, 16-CH3), 0.97 (3H, d,= 7.0 Hz, 18-CH3), 0.91 (1H, m, H-14), 0.70 (1H, dd,= 15.0, 8.3 Hz, H-13)；13C-NMR (125 MHz, CDCl3): 206.9 (C-9), 171.3 (20-OCOCH3), 138.9 (C-6), 136.8 (C-2), 130.2 (C-1), 128.7 (C-7), 84.5 (C-4), 80.8 (C-3), 73.9 (C-5), 72.7 (C-10), 66.8 (C-20), 44.3 (C-8), 40.0 (C-11), 31.1 (C-12), 28.6 (C-13), 24.1 (C-15), 23.3 (C-16), 23.1 (C-14), 21.3 (20-OCOCH3), 17.5 (C-18), 15.6 (C-19), 15.5 (C-17)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[24]，故鑒定化合物19為20--acetylingenol。

化合物20：無色油狀物，ESI-MS/: 377 [M＋H]+，分子式為C22H32O5。1H-NMR (500 MHz, CDCl3): 6.59 (1H, d,= 11.5 Hz, H-12), 6.15 (1H, d,= 11.1 Hz, H-5), 4.06 (1H, dd,= 10.8, 2.7 Hz, H-7), 3.86 (1H, dd,= 6.3, 3.7 Hz, H-3), 2.72 (1H, dd,= 14.7, 7.7 Hz, H-1), 2.65 (1H, dd,= 10.9, 6.6 Hz, H-4), 2.40 (1H, d,= 13.6 Hz, H-8), 2.27 (1H, m, H-1), 2.20 (1H, dd,= 7.0, 3.2 Hz, H-2), 2.05 (3H, s, 15-OCOCH3), 1.86 (3H, s, 20-CH3), 1.72 (1H, m, H-8), 1.51 (3H, s, 17-CH3), 1.44 (1H, dd,= 11.3, 8.3 Hz, H-11), 1.21 (3H, s, 18-CH3), 1.16 (1H, m, H-9), 1.12 (3H, d,= 8.9 Hz, 16-CH3), 1.11 (3H, s, 19-CH3)；13C-NMR (125 MHz, CDCl3): 194.8 (C-14), 169.8 (15-OCOCH3), 146.4 (C-6), 145.7 (C-12), 133.1 (C-13), 120.3 (C-5), 96.2 (C-15), 82.1 (C-3), 75.4 (C-7), 48.9 (C-4), 41.2 (C-2), 41.1 (C-1), 36.7 (C-8), 30.8 (C-9), 29.7 (C-11), 29.2 (C-18), 24.6 (C-10), 21.8 (15-OCOCH3), 19.1 (C-17), 18.5 (C-16), 16.4 (C-19), 12.4 (C-20)。以上數(shù)據(jù)與文獻(xiàn)報(bào)道一致[25]，故鑒定化合物20為altotibetol。

4 抗炎活性篩選

LPS可以刺激小鼠單核巨噬細(xì)胞RAW264.7生成誘導(dǎo)型一氧化氮合成酶（induced nitric oxide synthase，iNOS），進(jìn)而產(chǎn)生炎癥因子一氧化氮（nitric oxide，NO）。將RAW264.7細(xì)胞接種至96孔板，用1 μg/mL LPS進(jìn)行誘導(dǎo)刺激，同時(shí)加入待測(cè)化合物（終濃度50 μmol/L），同時(shí)設(shè)對(duì)照組（不含藥物）和陽性對(duì)照組（-NMMA）。細(xì)胞過夜培養(yǎng)后，取培養(yǎng)基檢測(cè)吸取培養(yǎng)基，通過Griess法在570 nm波長(zhǎng)測(cè)吸光度（）值來檢測(cè)亞硝酸鹽（NO2?），根據(jù)公式計(jì)算NO生成抑制率。在剩余培養(yǎng)基中加入MTS進(jìn)行細(xì)胞存活率檢測(cè)，排除化合物對(duì)細(xì)胞的毒性影響。活性測(cè)試結(jié)果見表1。

NO生成抑制率＝(對(duì)照－樣品)/對(duì)照

5 討論

對(duì)澤漆醋酸乙酯部位的研究中，共分離得到20個(gè)二萜類成分，包括10個(gè)假白欖烷二萜（1～10）、7個(gè)松香烷二萜（11～17）、1個(gè)阿替斯烷二萜（18）、1個(gè)巨大戟烷二萜（19）、1個(gè)續(xù)隨子烷二萜（20）。

經(jīng)文獻(xiàn)查閱，大戟屬的二萜通常具有細(xì)胞毒、抗炎等活性[2,14-15]。NO具有廣泛而重要的生物學(xué)調(diào)控功能，在炎癥、腫瘤及心血管系統(tǒng)等均有重要作用。當(dāng)免疫細(xì)胞遭受微生物內(nèi)毒素、炎癥介質(zhì)等刺激時(shí)，會(huì)生成大量的iNOS，產(chǎn)生NO進(jìn)行免疫應(yīng)答，因此抑制NO生成是化合物抗炎活性的直接指標(biāo)。因此，本研究測(cè)試了化合物1～20的NO生成抑制活性，結(jié)果顯示結(jié)構(gòu)骨架為賈白欖烷二萜的化合物6、8～10和19顯示了微弱的抗炎活性，說明賈白欖烷二萜是澤漆作為抗炎藥物的主要藥效物質(zhì)基礎(chǔ)。本研究為更好地開發(fā)利用澤漆奠定了一定的理論基礎(chǔ)。

表1 化合物1～20的NO生成抑制率(濃度50 μmol·L?1)

利益沖突 所有作者均聲明不存在利益沖突

[1] 秦友沐, 吳艷萍, 歐杜哈, 等. 澤漆化學(xué)成分研究[J].中草藥, 2018, 49(7): 1520-1524.

[2] Su J C, Cheng W, Song J G,. Macrocyclic diterpenoids fromand their potential anti-inflammatory activity [J]., 2019, 82(10): 2818-2827.

[3] 查顯進(jìn), 石強(qiáng), 邵峰, 等. 澤漆化學(xué)成分研究[J]. 中草藥, 2021, 52(2): 341-348.

[4] Schmidt R J, Evans F J. Skin irritants of the Sun spurge (L.) [J]., 1980, 6(3): 204-210.

[5] Nazir M, Ahmad W, Rabi N A,. Epicuticular leaf wax ofL. (Euphorbiaceae) [J]., 1993, 48(1/2): 5-9.

[6] Abdulladzhanova N G, Mavlyanov S M, Dalimov D N. Polyphenols of certain plants of the Euphorbiaceae family [J]., 2003, 39(4): 399-400.

[7] 姚學(xué)軍, 孟素蕊, 王喆. 澤漆的化學(xué)成分及其抗腫瘤轉(zhuǎn)移活性研究 [J]. 現(xiàn)代藥物與臨床, 2013, 28(6): 826-829.

[8] 楊莉, 陳海霞, 高文遠(yuǎn). 澤漆化學(xué)成分及藥理作用研究進(jìn)展 [J]. 中草藥, 2007, 38(10): 1585-1589.

[9] 陳學(xué)文. 澤漆粗提物對(duì)常見植物病原菌抑菌作用的初步研究 [J]. 浙江農(nóng)業(yè)科學(xué), 2005: 218-219.

[10] 何江波, 劉光明. 澤漆化學(xué)成分的初步研究 [J]. 大理學(xué)院學(xué)報(bào), 2010, 9(6): 5-7.

[11] 李娟, 濮社班. 大戟屬二萜類化學(xué)成分和生物活性研究進(jìn)展 [J]. 中國(guó)野生植物資源, 2017, 36(6): 36-44.

[12] 楊舜伊, 袁純紅, 陳業(yè)高. 大戟屬植物二萜化學(xué)成分和藥理活性研究新進(jìn)展 [J]. 中國(guó)野生植物資源, 2020, 39(6): 53-60.

[13] 史海明, 閔知大, 屠鵬飛, 等. 中國(guó)大戟屬植物中二萜成分的化學(xué)及生物活性 [J]. 化學(xué)進(jìn)展, 2008, 20(S1): 375-385.

[14] Liu C, Liao Z X, Liu S J,. Two new diterpene derivatives fromlunulata Bge and their anti-proliferative activities [J]., 2014, 96: 33-38.

[15] Tao H W, Hao X J, Liu P P,. Cytotoxic macrocyclic diterpenoids from[J]., 2008, 31(12): 1547-1551.

[16] Yamamura S, Shizuri Y, Kosemura S,. Diterpenes from[J]., 1989, 28(12): 3421-3436.

[17] Chen H Q, Wang H, Yang B,. Diterpenes inhibiting NO production from[J]., 2014, 95: 133-138.

[18] Lu Z Q, Guan S H, Li X N,. Cytotoxic diterpenoids from[J]., 2008, 71(5): 873-876.

[19] Mai Z P, Ni G, Liu Y F,. Helioscopianoids A-Q, bioactive jatrophane diterpenoid esters from[J]., 2018, 8(5): 805-817.

[20] Zhang W, Guo Y W. Chemical studies on the constituents of the Chinese medicinal herbL [J].(), 2006, 54(7): 1037-1039.

[21] Borghi D, Baumer L, Ballabio M,. Structure elucidation of helioscopinolides D and E fromcell cultures [J]., 1991, 54(6): 1503-1508.

[22] Crespi-Perellino N, Garofano L, Arlandini E,. Identification of new diterpenoids fromcell cultures [J]., 1996, 59(8): 773-776.

[23] Lal A R, Cambie R C, Rutledge P S,. Ent-atisane diterpenes from[J]., 1990, 29(6): 1925-1935.

[24] Meng X H, Wang K, Chai T,. Ingenane and jatrophane diterpenoids fromand their antiproliferative effects [J]., 2020, 172: 112257.

[25] Zhang Z X, Qi F M, Li H H, Dong L L, Hai Y, Fan G X, Fei D Q. A new lathyrane diterpenoid from the whole plant of[J]., 2014, 35(2): 641-643.

Diterpenoids from whole herb of

WANG Yan, LIANG Xu-bo, ZHAO Zhen-zhu

School of Pharmacy, Henan University of Chinese Medicine, Zhengzhou 450046, China

To investigate the diterpenoids from the whole herb of.The ethyl acetate extract ofextracted by EtOH (95%) was separated and purified by various chromatographic columns, including macroporous resins, silica gels, medium pressure liquid chromatography (MPLC), Sephadex LH-20, and preparative high performance liquid chromatography (prep-HPLC). The structures of purified compounds were determined by physiochemical properties and spectroscopic data.Twenty diterpenoids were isolated from of, and they are identified as euphoscopin A (1), euphoscopin B (2), euphoscopin C (3), euphoscopin E (4), euphorbiapene D (5), euphornin A (6), euphornin B (7), euphornin (8), helioscopianoid M (9), euphoheliosnoid D (10), 2α-hydroxy helioscopinolide B (11), helioscopinolide A (12), helioscopinolide B (13), helioscopinolide C (14), helioscopinolide D (15), helioscopinolide H (16), helioscopinolide L (17),-16β,17-dihydroxyatlsan-3-one (18), 20--acetylingenol (19) and altotibetol (20). Additionally, all compounds were tested for their inhibitory activity against NO production.All compounds were isolated from different plants ofbefore, among which compounds 15, 16, 17, and 20 are isolated fromfor the first time. And compounds 6, 8—10 and 19 showed weak anti-inflammatory activity.

L.; diterpenoids; anti-inflammatory activity; euphornin A; helioscopinolide D; altotibetol

R284.1

A

0253 - 2670(2022)15 - 4625 - 09

10.7501/j.issn.0253-2670.2022.15.005

2021-12-23

國(guó)家自然科學(xué)基金資助項(xiàng)目（82003607）；河南省高等學(xué)校重點(diǎn)科研項(xiàng)目（21A360002）；河南省中醫(yī)藥科學(xué)研究專項(xiàng)（20-21ZY1039）

王 妍，女，碩士研究生，從事中藥活性成分研究。E-mail: k921_w328@163.com

通信作者：梁旭博，男，碩士，從事中藥活性成分研究。E-mail: 15838258630@163.com

趙珍珠，女，博士，講師，從事中藥活性成分研究。E-mail: zzhenzhu0921@163.com

[責(zé)任編輯 王文倩]