徐蓮蓮，陳志超，何夢(mèng)麗，楊 寶，付劍江，陳芳有，羅永明

?化學(xué)成分 ?

卡氏乳香中1個(gè)新的降二萜

徐蓮蓮，陳志超，何夢(mèng)麗，楊 寶，付劍江，陳芳有*，羅永明*

江西中醫(yī)藥大學(xué)藥學(xué)院，江西 南昌 330004

研究卡氏乳香中的化學(xué)成分及其抗骨質(zhì)疏松活性。運(yùn)用硅膠柱色譜、ODS柱色譜和制備高效液相色譜等方法進(jìn)行化學(xué)成分分離，通過(guò)波譜數(shù)據(jù)及文獻(xiàn)對(duì)比對(duì)其進(jìn)行結(jié)構(gòu)解析。卡氏乳香的95%乙醇提取物中分離得到11個(gè)化合物，分別鑒定為 (1,2,8,9)-1,9-環(huán)氧-8-乙酰氧基-4,5-二氧代降西松烷型-2-烯（1）、(1,3,7,11,12)-1,12-epoxy-4-methylenecembr-7-ene-3,11-diol（2）、boscartin F（3）、boscartene K（4）、fidmansumbin-13(17)-en-3,16-diene（5）、3α-acetoxyl-7-oxo-tirucalla-8,24-dien-21-oic acid（6）、boscartene N（7）、3β-hydroxy-tirucallic acid（8）、α-boswellic acid（9）、boscarterol O（10）、boscarterol F（11）?；衔?是新化合物，命名為卡氏乳香素，化合物5為首次從乳香中分離得到，化合物2和10首次從該屬植物中分離得到。對(duì)11個(gè)化合物進(jìn)行抗骨質(zhì)疏松活性篩選，發(fā)現(xiàn)化合物1、3、6和10具有良好的抑制破骨細(xì)胞活性。

卡氏乳香；萜類(lèi)化合物；降二萜；(1,2,8,9)-1,9-環(huán)氧-8-乙酰氧基-4,5-二氧代西松烷型-2-烯；卡氏乳香素

乳香是橄欖科植物乳香樹(shù)Birdw.及同屬植物Birdw.樹(shù)皮滲出的膠狀樹(shù)脂，具有活血定痛、消腫生肌的功效[1]。大量的臨床試驗(yàn)結(jié)果表明，乳香具有較好的抗炎活性，主要用于治療類(lèi)風(fēng)濕性關(guān)節(jié)炎、潰瘍性結(jié)腸炎和骨關(guān)節(jié)炎[2-3]。乳香屬大約包含25種，其中卡氏乳香.Birdw.受到研究人員廣泛的關(guān)注，研究表明其主要化學(xué)成分為三萜類(lèi)和二萜類(lèi)[4]。早期文獻(xiàn)研究主要集中于乳香酸等三萜類(lèi)成分，近年來(lái)從中報(bào)道了一些結(jié)構(gòu)新穎、活性多樣的西松烷型二萜，但研究尚不深入[5]。本研究對(duì)卡氏乳香95%乙醇提取物的化學(xué)成分進(jìn)行研究，從中分離鑒定出11個(gè)化合物，分別為 (1,2,8,9)-1,9-環(huán)氧-8-乙酰氧基-4,5-二氧代西松烷型-2-烯[(1,2,8,9)-1,9-epoxy-8-acetoxy-4,5-dioxo-2-cembranene，1]、(1,3,7,11,12)-1,12-epoxy-4- methylenecembr-7-ene-3,11-diol（2）、boscartin F（3）、boscartene K（4）、fidmansumbin-13(17)-en- 3,16-diene（5）、3α-acetoxyl-7-oxo-tirucalla-8,24- dien-21-oic acid（6）、boscartene N（7）、3β-hydroxy- tirucallic acid（8）、α-boswellic acid（9）、boscarterol O（10）、boscarterol F（11）。其中化合物1是新化合物，命名為卡氏乳香素，化合物5為首次從乳香中分離得到，化合物2和10首次從該屬植物中分離得到。

1 儀器與材料

Bruker Avance Ⅲ HD 600 MHz型核磁共振波譜儀（Bruker公司，瑞士）；AB SCIEX Triple ESI 5600+型高分辨飛行時(shí)間質(zhì)譜聯(lián)用儀（AB SCIEX公司，美國(guó)）；Jasco J-1500型圓二色譜儀（Jasco公司，日本）；Rulph Autopol IV自動(dòng)旋光儀（Rudolph Research Analytical公司，美國(guó)）；Agilent 1260型高效液相色譜儀（Agilent公司，美國(guó)）；FTIR-650紅外光譜儀（天津港東公司）；Waters 515 制備型液相色譜儀（Waters公司，美國(guó)）；C18半制備色譜分析柱（250 mm×10 mm，5 μm，YMC，日本）；柱色譜硅膠（200～300目）和薄層色譜硅膠GF254（青島海洋化工廠）；色譜純甲醇（西隴科學(xué)股份有限公司）；純凈水（杭州娃哈哈集團(tuán)有限公司）；分析純石油醚（PE）、二氯甲烷、醋酸乙酯和甲醇（西隴科學(xué)股份有限公司）。

卡氏乳香藥材（批號(hào)20190401）于2019年10月購(gòu)自于江西繼中堂健康科技有限公司，經(jīng)南昌大學(xué)第一附屬醫(yī)院周健副主任藥師鑒定為卡氏乳香樹(shù)Birdw.樹(shù)皮滲出的膠狀樹(shù)脂。樣品（20191020）保存于江西中醫(yī)藥大學(xué)藥學(xué)院中藥化學(xué)教研室。

2 提取與分離

卡氏乳香樹(shù)脂10 kg，粉碎后用95%乙醇冷浸提取3次，合并提取液減壓濃縮，得到總浸膏6.7 kg，所得浸膏經(jīng)硅藻土柱色譜依次用石油醚、二氯甲烷、醋酸乙酯和甲醇進(jìn)行分離，回收有機(jī)溶劑，分別得到石油醚部位2.9 kg、二氯甲烷部位1.8 kg、醋酸乙酯部位850.0 g和甲醇部位200.0 g。二氯甲烷部位浸膏經(jīng)PRP 512A樹(shù)脂洗脫分離（30%、50%、70%、95%乙醇-水溶液）得到4個(gè)部分。

取70%乙醇組分（400.0 g）經(jīng)硅膠柱色譜分離，以石油醚-醋酸乙酯（30∶1→0∶1）梯度洗脫得到7個(gè)組分（Fr. 1～7），其中Fr. 4（184.0 g）經(jīng)減壓柱色譜以二氯甲烷-甲醇（100∶1→0∶1）得到6個(gè)組分（Fr. 4-1～4-6）。Fr. 4-1（50.0 g）經(jīng)MCI柱色譜洗脫（50%→100%甲醇-水溶液）得到6個(gè)組分（Fr. 4-1-1～4-1-6)，其中Fr. 4-1-6（17.0 g）進(jìn)行反復(fù)硅膠柱色譜分離得到化合物6（600.0 mg）、8（202.0 mg）、9（29.1 mg）和11（33.0 mg），F(xiàn)r. 4-1-5（30.0 g）經(jīng)反復(fù)硅膠柱色譜和制備液相色譜（80%甲醇-水溶液，體積流量2 mL/min）分離純化得到化合物4（7.0 mg，R＝1.2 min）、5（13.1 mg，R＝1.5 min）和7（6.2 mg，R＝1.8 min）。對(duì)Fr. 4-3進(jìn)行ODS柱色譜（洗脫梯度為60%～100%的甲醇-水溶液）得到5個(gè)組分（Fr. 4-3-1～4-3-5），其中Fr. 4-3-1經(jīng)反復(fù)硅膠柱色譜和制備液相柱色譜（65%甲醇-水溶液，體積流量2 mL/min）分離得到化合物1（5.6 mg，R＝2.0 min）、2（11.1 mg，R＝2.1 min）和3（9.6 mg，R＝1.8 min）。Fr. 4-6（14.0 g）經(jīng)ODS柱色譜（60%→100%甲醇-水溶液）進(jìn)行梯度洗脫得到5個(gè)組分（Fr. 4-6-1～4-6-5），其中Fr. 4-6-1經(jīng)制備液相柱色譜（60%甲醇-水溶液，體積流量2 mL/min）得到化合物10（12.2 mg，R＝2.2 min）。

3 結(jié)構(gòu)鑒定

通過(guò)1H-1H COSY譜和HMBC（圖1）可以對(duì)以上做進(jìn)一步證實(shí)。1H-1H COSY譜顯示H-2/H-3、H-6/H-7/H-8、H-10/H-11和H-12/H3-13/H3-14相關(guān)信號(hào)。在HMBC譜中，H-2 (H6.75) 與C-1、C-3、C-4和C-11相關(guān)，且H-3 (H6.24) 與C-1、C-2、C-4和C-15相關(guān)，H-15 (H2.30) 與C-3和C-4相關(guān)，由此確定C-2、C-3、C-4和C-15為α, β不飽和酮部分；H-12 (H1.79) 與C-1、C-2、C-11、C-13和C-14相關(guān)，H-17 (H1.20) 與C-8、C-9和C-10相關(guān)，結(jié)合C-1和C-9的化學(xué)位移可以確定C-1和C-9之間連有1個(gè)五元氧環(huán)[6]；H-16 (H2.14)與C-5和C-6相關(guān)，H-6a (H2.49) 與C-5和C-7相關(guān)，可以確定C-5為酮羰基部分；H-13 (H0.91) 和C-1、C-12和C-14相關(guān), H-14 (H0.87) 與C-1、C-12和C-13相關(guān)，H-12 (H1.79) 與C-1、C-12、C-13相關(guān)，可以確定C-1上連有1個(gè)異丙基結(jié)構(gòu)。

在NOESY譜中，觀察到H-17與H8/H12的相關(guān)信號(hào)，表明H-8/H-12/H-17位于同一平面內(nèi)?；衔?的ECD圖譜在234 nm (= ?4.70) 處呈現(xiàn)負(fù)的Cotton效應(yīng)，化合物1的計(jì)算ECD和實(shí)驗(yàn)ECD圖譜基本一致（圖2），確定該化合物的絕對(duì)構(gòu)型為1,8,9,其結(jié)構(gòu)如圖1所示。通過(guò)以上信息推測(cè)化合物1的結(jié)構(gòu)為 (1,2,8,9)-1,9-環(huán)氧-8-乙酰氧基-4,5-二氧代西松烷型-2-烯，命名為卡氏乳香素。波譜數(shù)據(jù)歸屬見(jiàn)表1。

化合物2：白色針狀結(jié)晶（甲醇）；HR-ESI-MS/323.255 0 [M＋H]+（C20H35O3計(jì)算值為323.258 6）。1H-NMR (600 MHz, CDCl3): 5.33 (1H, t,= 7.5 Hz, H-7), 4.99 (1H, s, H-18b), 4.89 (1H, s, H-18a), 4.28 (1H, d,= 9.9 Hz, H-3), 3.38 (1H, d,= 9.5 Hz, H-11), 2.37 (1H, ddd,= 13.1, 9.9, 3.1 Hz, H-5a), 2.28 (1H, m, H-15), 2.23 (1H, ddd,=10.0, 7.3, 3.1 Hz, H-6a), 2.17 (1H, m, H-9a), 2.08 (1H, m, H-5b), 2.04 (1H, m, H-9b), 2.02 (1H, m, H-14a), 2.00 (1H, m, H-13a), 1.91 (1H, m, H-10a), 1.78 (1H, m, H-13b), 1.72 (1H, dd,= 14.6, 9.9 Hz, H-2a), 1.67 (3H, s, H-19), 1.61 (1H, dd,= 14.6, 1.5 Hz, H-2b), 1.50 (1H, m, H-14b), 1.46 (1H, m, H-6b), 1.42 (1H, m, 10a), 1.12 (3H, s, H-20), 0.90 (3H, d,= 2.5 Hz, H-16), 0.89 (3H, d,= 2.6 Hz, H-17)；13C-NMR (150 MHz, CDCl3): 88.9 (C-1), 40.6 (C-2), 70.0 (C-3), 155.1 (C-4), 34.1 (C-5), 29.9 (C-6), 126.5 (C-7), 135.0 (C-8), 33.4 (C-9), 31.2 (C-10), 76.2 (C-11), 85.1 (C-12), 36.2 (C-13), 31.8 (C-14), 33.4 (C-15), 18.2 (C-16), 17.4 (C-17), 110.0 (C-18), 18.6 (C-19), 20.2 (C-20)。以上數(shù)據(jù)與文獻(xiàn)對(duì)照基本一致[7]。對(duì)化合物2進(jìn)行X-射線單晶衍射分析，確定其絕對(duì)構(gòu)型，見(jiàn)圖3。晶體數(shù)據(jù)：分子式為C20H34O3,＝323.499，斜方晶系，晶胞參數(shù)＝0.646 71 (3) nm，＝1.912 80 (9) nm，＝0.790 44 (3) nm，＝0. 947 83 (7) nm3，＝2，＝293（2），Kμ (Cu K)＝0.577 mm?1，空間集群為P212121(No. 19), 收集衍射點(diǎn)數(shù)為7629，獨(dú)立衍射點(diǎn)數(shù)為3193，int＝0.044 4，sigma＝0.054 1，1＝0.059 5，2＝0.146 3，F(xiàn)lack常數(shù)為?0.1 (2)。因此，化合物1確定為(13,7,11,12)-1,12-epoxy-4- methylenecembr-7-ene-3,11-diol。

圖1 化合物1的主要1H-1H COSY、HMBC和NOESY相關(guān)

圖2 化合物1的實(shí)驗(yàn)和計(jì)算ECD圖

表1 化合物1的1H-NMR和13C-NMR數(shù)據(jù)(600/150MHz, CDCl3)

Table1 1H-NMR and 13C-NMR data of compound 1 (600/150MHz, CDCl3)

碳位δHδc 1 89.2 26.75 (1H, d, J = 16.0 Hz)149.9 36.24 (1H, d, J = 16.0 Hz)129.3 4 198.7 5 208.0 62.41 (1H, m), 2.49 (1H, m)40.2 71.72 (1H, m), 1.76 (1H, m)24.2 84.79 (1H, dd, J = 10.5, 2.6 Hz)77.9 9 84.6 101.65 (1H, m), 1.76 (1H, m)35.0 111.94 (1H, m), 2.00 (1H, m)33.2 121.79 (1H, dd, J = 6.8, 7.0 Hz)38.3 130.91 (1H, d, J = 7.0 Hz)17.8 140.87 (1H, d, J = 6.8 Hz)18.4 152.30 (3H, s)28.2 162.14 (3H, s)30.2 171.20 (3H, s)22.4 18 171.0 192.02 (3H, s)21.3

圖3 化合物2的橢球圖(橢球率為50%)

化合物3：白色針狀結(jié)晶（甲醇）；HR-ESI-MS/337.235 9 [M＋H]+（C20H33O4計(jì)算值為337.237 9）。1H-NMR (600 MHz, CDCl3): 6.84 (1H, dd,= 6.3, 5.8 Hz, H-7), 3.86 (1H, d,= 12.5 Hz, H-10a), 3.53 (1H, d,= 10.4 Hz, H-11), 3.14 (1H, dd,= 4.7, 2.3 Hz, H-3), 2.46 (1H, m, H-6a), 2.46 (1H, m, H-6b), 2.27 (1H, m, H-5a), 2.21 (1H, sept,= 6.8 Hz, H-15), 2.11 (1H, m, H-13a), 2.05 (1H, dd,= 12.5, 10.4 Hz, H-10b), 2.02 (1H, m, H-14a), 1.87 (1H, m, H-13b), 1.82 (1H, dd,= 15.8, 2.3 Hz, H-2a), 1.78 (3H, s, H-19), 1.62 (1H, dd,= 15.8, 4.7 Hz, H-2b), 1.48 (1H, m, H-14b), 1.36 (1H, m, H-5b), 1.14 (3H, s, H-20), 1.13 (3H, s, H-18), 0.98 (3H, d,= 6.9 Hz, H-17), 0.92 (3H, d,= 6.9 Hz, H-16)；13C-NMR (150 MHz, CDCl3): 89.0 (C-1), 37.4 (C-2), 60.8 (C-3), 58.7 (C-4), 37.9 (C-5), 25.6 (C-6), 144.5 (C-7), 137.2(C-8), 203.5 (C-9), 40.9 (C-10), 78.3 (C-11), 84.9 (C-12), 36.8 (C-13), 30.5 (C-14), 33.0 (C-15), 18.8 (C-16), 17.0 (C-17), 16.3 (C-18), 11.4 (C-19), 19.8 (C-20)。以上數(shù)據(jù)與文獻(xiàn)對(duì)照基本一致[8]，故鑒定該化合物為boscartin F。

化合物4：白色針狀結(jié)晶（甲醇）；HR-ESI-MS/373.272 9 [M＋H]+（C24H37O3計(jì)算值為373.274 3）。1H-NMR (600 MHz, CDCl3): 5.80 (1H, d,= 1.3 Hz, H-17), 4.68 (1H, t,= 2.6 Hz, H-3), 2.71 (1H, dd,= 14.2, 4.7 Hz, H-12a), 2.56 (1H, d,= 19.1 Hz, H-15a), 2.42 (1H, td,= 13.6, 6.8 Hz, H-12b), 2.10 (3H, s, CH3CO-), 1.94 (1H, m, H-2a), 1.88 (1H, d,= 19.1 Hz, H-15b), 1.84 (1H, m, H-11a), 1.70 (1H, m, H-9), 1.67 (1H, m, H-7a), 1.64 (1H, m, H-2b), 1.54 (1H, m, H-1a), 1.53 (1H, m, H-11b), 1.48 (1H, m, H-6a), 1.40 (1H, m, H-6b), 1.35 (3H, s, H-22), 1.31 (1H, m, H-7b), 1.28 (1H, m, H-1a), 1.28 (1H, m, H-5), 0.89 (3H, s, H-20), 0.88 (3H, s, H-19), 0.87 (3H, s, H-21), 0.80 (3H, s, H-18)；13C-NMR (150 MHz, CDCl3): 34.4 (C-1), 23.1 (C-2), 78.1 (C-3), 36.9 (C-4), 50.8 (C-5), 18.2 (C-6), 35.5 (C-7), 41.5 (C-8), 50.2 (C-9), 37.7 (C-10), 22.1 (C-11), 27.5 (C-12), 190.6 (C-13), 51.6 (C-14), 46.2 (C-15), 209.5 (C-16), 126.6 (C-17), 16.1 (C-18), 16.3 (C-19), 22.0 (C-20), 28.0 (C-21), 22.6 (C-22), 170.9 (CH3CO-), 21.6 (CH3CO-)。以上數(shù)據(jù)與文獻(xiàn)對(duì)照基本一致[9]，故鑒定該化合物為boscartene K。

化合物5：黃色油狀物；HR-ESI-MS/329.248 1 [M＋H]+（C22H33O2計(jì)算值為329.248 1）。1H-NMR (600 MHz, CDCl3): 5.79 (1H, s, H-17), 2.71 (1H, ddd,= 14.1, 5.0, 1.9 Hz, H-12a)，2.53 (1H, d,= 19.1 Hz, H-15a), 2.49 (1H, m, H-2a), 2.49 (1H, m, H-2b), 2.43 (1H, m, H-12b), 1.99 (1H, m, H-1a), 1.88 (1H, d,= 19.1 Hz, H-15b), 1.80 (1H, m, H-11a), 1.66 (1H, dd,= 12.7, 3.1 Hz, H-9), 1.66 (1H, m, H-6a), 1.55 (1H, m, H-7a), 1.51 (1H, m, H-1b), 1.46 (1H, m, H-6b), 1.42 (1H, m, H-7b), 1.42 (1H, m, H-11b), 1.31 (3H, s, H-22), 1.27 (1H, m, H-5), 1.11 (3H, s, H-20), 1.05 (3H, s, H-21), 0.96 (3H, s, H-19), 0.84 (3H, d,= 0.8 Hz, H-18)；13C-NMR (150 MHz, CDCl3): 39.8 (C-1), 34.1 (C-2), 217.5 (C-3), 47.4 (C-4), 55.2 (C-5), 19.7 (C-6), 50.1 (C-7), 41.1 (C-8), 51.1 (C-9), 37.3 (C-10), 22.4 (C-11), 27.5 (C-12), 187.4 (C-13), 51.5 (C-14), 46.1 (C-15), 209.2 (C-16), 126.9 (C-17), 15.9 (C-18), 16.4 (C-19), 26.9 (C-20), 21.0 (C-21), 22.7 (C-22)。以上數(shù)據(jù)與文獻(xiàn)對(duì)照基本一致[10]，故鑒定該化合物為fidmansumbin-13(17)-en- 3,16-diene。

化合物6：白色粉末；HR-ESI-MS/513.357 4 [M＋H]+（C32H49O5計(jì)算值為513.358 0）。1H-NMR (600 MHz, DMSO-6): 11.60 (1H, s, COOH), 5.09 (1H, t,= 6.4 Hz, H-24), 4.57 (1H, s, H-3), 2.39 (1H, m, H-2a), 2.36 (1H, m, H-2b), 2.34 (1H, m, H-6b), 2.32 (1H, m, H-6a), 2.27 (1H, m, H-11a), 2.22 (1H, m, H-11b), 2.18 (1H, m, H-16b), 2.10 (1H, m, H-5), 2.03 (1H, m, H-17), 2.02 (3H, s, CH3CO-), 2.00 (1H, m, H-15a), 1.94 (1H, m, H-16a), 1.94 (1H, m, H-23a), 1.90 (1H, m, H-23b), 1.72 (1H, m, H-1a), 1.59 (3H, s, H-26), 1.54 (3H, s, H-27), 1.45 (1H, m, H-12a), 1.45 (1H, m, H-15b), 1.22 (1H, m, H-22a), 1.22 (1H, m, H-22b), 1.17 (1H, m, H-1b), 1.12 (1H, m, H-12b), 1.06 (1H, m, H-16b), 1.03 (3H, s, H-19), 0.97 (3H, s, H-29), 0.96 (3H, s, H-28), 0.89 (3H, s, H-30), 0.80 (3H, s, H-18)；13C-NMR (150 MHz, DMSO-6): 32.2 (C-1), 22.4 (C-2), 76.1 (C-3), 36.2 (C-4), 47.3 (C-5), 34.9 (C-6), 196.6 (C-7), 137.5 (C-8), 165.6 (C-9), 38.7 (C-10), 21.1 (C-11), 27.6 (C-12), 43.1 (C-13), 43.9 (C-14), 29.0 (C-15), 26.9 (C-16), 46.8 (C-17), 15.7 (C-18), 17.9 (C-19), 45.4 (C-20), 177.1 (C-21), 31.0 (C-22), 25.5 (C-23), 123.8 (C-24), 131.2 (C-25), 22.7 (C-26), 17.5 (C-27), 25.5 (C-28), 26.6 (C-29), 24.1 (C-30), 169.9 (CH3CO-), 20.9 (CH3CO-)。以上數(shù)據(jù)與文獻(xiàn)對(duì)照基本一致[11]，故鑒定該化合物為3α- acetoxyl-7-oxo-tirucalla-8,24- dien-21-oic acid。

化合物7：白色粉末；HR-ESI-MS/467.313 3 [M＋H]+（C30H43O4計(jì)算值為467.316 7）。1H-NMR (600 MHz, CDCl3): 5.22 (1H, m, H-24), 5.20 (1H, m, H-23), 2.78 (1H, ddd,= 15.4, 13.8, 6.0 Hz, H-2), 2.68 (1H, td,= 8.9, 4.9 Hz, H-20), 2.49 (1H, d,= 13.9 Hz, H-6a), 2.43 (1H, m, H-6b), 2.41 (1H, m, H-2b), 2.40 (1H, m, H-11), 2.38 (1H, m, H-22a), 2.29 (1H, m, H-17), 2.23 (1H, m, H-15a), 2.18 (1H, m, H-1a), 2.13 (1H, dd,= 13.8, 4.3 Hz, H-5), 2.04 (1H, m, H-12a), 1.99 (1H, m, H-16a), 1.78 (1H, m, H-1b), 1.78 (1H, m, H-12b), 1.76 (3H, s, H-26), 1.75 (3H, s, H-27), 1.66 (1H, m, H-15b), 1.46 (1H, m, H-16b), 1.29 (3H, s, H-19), 1.13 (3H, s, H-29), 1.09 (3H, s, H-28), 1.03 (3H, s, H-30), 0.83 (3H, s, H-18)；13C-NMR (150 MHz, CDCl3): 35.5 (C-1), 34.6 (C-2), 214.5 (C-3), 47.5 (C-4), 49.5 (C-5), 36.3 (C-6), 197.5 (C-7), 139.1 (C-8), 164.7 (C-9), 39.3 (C-10), 24.1 (C-11), 27.7 (C-12), 47.5 (C-13), 44.9 (C-14), 31.3 (C-15), 24.7 (C-16), 43.3 (C-17), 16.7 (C-18), 18.2 (C-19), 41.8 (C-20), 178.9 (C-21), 35.0 (C-22), 75.3 (C-23), 123.2 (C-24), 139.1 (C-25), 25.9 (C-26), 18.6 (C-27), 24.1 (C-28), 21.6 (C-29), 24.7 (C-30)。以上數(shù)據(jù)與文獻(xiàn)對(duì)照基本一致[12]，故鑒定該化合物為boscartene N。

化合物8：白色粉末，HR-ESI-MS/457.367 8 [M＋H]+（C30H49O3計(jì)算值為457.368 2）。1H-NMR (600 MHz, CDCl3): 5.09 (1H, m, H-24), 3.23 (1H, dd,= 11.7, 4.5 Hz, H-3), 2.28 (1H, m, H-17), 2.08 (1H, m, H-20), 2.08 (1H, m, H-11a), 2.05 (1H, m, H-23a), 1.98 (1H, m, H-11b), 1.95 (1H, m, H-24b), 1.76 (1H, m, H-12a), 1.74 (1H, m, H-12b), 1.70 (1H, m, H-2a), 1.68 (3H, s, H-26), 1.66 (1H, m, H-1b), 1.58 (3H, s, H-27), 1.56 (2H, m, H-22), 1.56 (2H, m, H-7), 1.53 (1H, m, H-6a), 1.41 (1H, dd,= 12.4, 6.2 Hz, H-2b), 1.35 (2H, m, H-16), 1.35 (1H, m, H-6b), 1.25 (1H, m, H-15a), 1.22 (1H, m, H-15b), 1.18 (1H, m, H-1a), 1.12 (1H, dd,= 12.4, 1.9 Hz, H-5), 1.00 (3H, s, H-29), 0.93 (3H, s, H-17), 0.88 (3H, s, H-30), 0.82 (3H, s, H-28), 0.74 (3H, s, H-18)；13C-NMR (150 MHz, CDCl3): 35.4 (C-1), 27.7 (C-2), 79.1 (C-3), 39.1 (C-4), 51.1 (C-5), 19.0 (C-6), 29.0 (C-7) , 134.2 (C-8), 132.4 (C-9), 37.4 (C-10), 26.1 (C-11), 32.6 (C-12), 44.0 (C-13), 49.7 (C-14), 29.5 (C-15), 28.0 (C-16), 47.1 (C-17), 20.2 (C-18), 15.7 (C-19), 47.6 (C-20), 182.3 (C-21), 21.7 (C-22), 27.0 (C-23), 123.7 (C-24), 133.3 (C-25), 24.6 (C-26), 17.8 (C-27), 28.2 (C-28), 15.9 (C-29), 25.8 (C-30)。以上數(shù)據(jù)與文獻(xiàn)對(duì)照基本一致[13]，故鑒定該化合物為3βhydroxy-tirucallic acid。

化合物9：白色粉末，HR-ESI-MS/457.367 8 [M＋H]+（C30H49O3計(jì)算值為457.368 2）。1H-NMR (600 MHz, CDCl3): 5.19 (1H, t,= 3.6 Hz, H-12), 4.08 (1H, t,= 2.8 Hz, H-3), 2.21 (1H, m, H-11a), 1.99 (1H, m, H-11b), 1.95 (1H, dd,= 14.0, 4.8 Hz, H-18), 1.89 (1H, m, H-21), 1.88 (1H, m, H-22), 1.77 (1H, m, H-21b), 1.71 (1H, m, H-2a), 1.68 (1H, m, H-22b), 1.64 (1H, m, H-6a), 1.59 (1H, m, H-1a), 1.52 (1H, m, H-7a), 1.49 (1H, m, H-2b), 1.44 (1H, m, H-19a), 1.43 (1H, m, H-7b), 1.40 (1H, m, H-6b), 1.37 (1H, m, H-5), 1.35 (3H, s, H-23), 1.32 (1H, m, H-1b), 1.27 (1H, m, H-15a), 1.23 (1H, m, H-16a), 1.21 (1H, m, H-19b), 1.15 (3H, s, H-27), 1.10 (1H, m, H-15b), 1.03 (1H, m, H-9), 1.00 (3H, s, H-26), 0.98 (1H, m, H-16b), 0.88 (3H, s, H-25), 0.87 (3H, s, H-29), 0.87 (3H, s, H-30), 0.83 (3H, s, H-28)；13C-NMR (150 MHz, CDCl3): 33.5 (C-1), 26.1 (C-2), 70.6 (C-3), 47.3 (C-4), 49.0 (C-5), 19.6 (C-6), 32.7 (C-7), 39.7 (C-8), 46.7 (C-9), 37.5 (C-10), 23.6 (C-11), 121.6 (C-12), 145.0 (C-13), 41.8 (C-14), 26.1 (C-15), 26.8 (C-16), 32.7 (C-17), 47.3 (C-18), 46.7 (C-19), 31.0 (C-20), 34.6 (C-21), 37.5 (C-22), 24.0 (C-23), 183.2 (C-24), 13.0 (C-25), 16.6 (C-26), 26.1 (C-27), 28.3 (C-28), 33.5 (C-29), 23.6 (C-30)。以上數(shù)據(jù)與文獻(xiàn)對(duì)照基本一致[14]，故鑒定該化合物為α-boswellic acid。

化合物10：黃色油狀物，HR-ESI-MS/339.190 9 [M＋Na]+（C20H28NaO3計(jì)算值為339.193 6）。1H-NMR (600 MHz, CDCl3): 7.1 (1H, m, H-17), 4.82 (1H, d,= 2.1 Hz, H-16), 4.71 (1H, s, H-12b), 4.69 (1H, s, H-12a), 2.45 (1H, dd,= 13.3, 6.3 Hz, H-9a), 2.27 (1H, td,= 9.6, 6.6 Hz, H-1), 2.16 (1H, m, H-9b), 2.04 (1H, m, H-8a), 1.91 (1H, m, H-2a), 1.77 (3H, s, H-20), 1.75 (1H, m, H-3a), 1.68 (1H, m, H-3b), 1.65 (1H, m, H-2b), 1.65 (1H, m, H-15), 1.37 (1H, d,= 10.8 Hz, H-5), 1.34 (3H, s, H-11), 1.29 (1H, m, H-7), 1.07 (1H, m, H-8b), 1.04 (1H, m, H-6), 0.93 (3H, s, H-14)；13C-NMR (150 MHz, CDCl3): 51.7 (C-1), 25.9 (C-2), 41.6 (C-3), 81.2 (C-4), 53.3 (C-5), 25.9 (C-6), 24.4 (C-7), 25.0 (C-8), 38.7 (C-9), 153.6 (C-10), 25.2 (C-11), 106.7 (C-12), 23.3 (C-13), 12.0 (C-14), 65.8 (C-15), 72.5 (C-16), 155.6 (C-17), 143.6 (C-18), 207.1 (C-19), 10.2 (C-20)。以上數(shù)據(jù)與文獻(xiàn)對(duì)照基本一致[15]，故鑒定該化合物為boscarterol O。

化合物11：白色油狀物，HR-ESI-MS/301.212 1 [M＋H]+（C20H29O2計(jì)算值為301.216 8）。1H-NMR (600 MHz, CDCl3): 9.39 (1H, s, H-19), 6.81 (1H, d,= 11.2 Hz, H-15), 6.46 (1H, dd,=15.1, 11.2 Hz, H-16), 5.82 (1H, d,= 15.1 Hz, H-17), 4.75 (1H, s, H-12a), 4.73 (1H, s, H-12b), 2.48 (1H, dd,= 13.6, 6.2 Hz, H-9a), 2.26 (1H, td,= 10.6, 6.1 Hz, H-1), 2.07 (1H, t,= 1.8 Hz, H-9b), 1.95 (1H, dq,= 11.6, 6.4 Hz, H-2a), 1.95 (1H, m, H-8a), 1.85 (3H, s, H-20), 1.79 (1H, td,= 13.1, 6.3, 2.4 Hz, H-3a), 1.67 (1H, m, H-2b), 1.61 (1H, td,= 12.8, 11.5, 6.2 Hz, H-3b), 1.52 (1H, t,= 10.8 Hz, H-5), 1.29 (3H, s, H-14), 1.26 (1H, m, H-7), 1.24 (3H, s, H-11), 1.20 (1H, m, H-8b), 1.03 (1H, t,= 11.2, 9.7 Hz, H-6)；13C-NMR (150 MHz, CDCl3): 52.9 (C-1), 26.8 (C-2), 42.1 (C-3), 80.9 (C-4), 53.5 (C-5), 33.0 (C-6), 30.7 (C-7), 24.4 (C-8), 38.3 (C-9), 152.6 (C-10), 26.3 (C-11), 107.4 (C-12), 28.4 (C-13), 12.3 (C-14), 156.0 (C-15), 120.3 (C-16), 150.1 (C-17), 135.1 (C-18), 195.1 (C-19), 9.6 (C-20)。以上數(shù)據(jù)與文獻(xiàn)對(duì)照基本一致[16]，故鑒定該化合物為boscarterol F。

4 抗骨質(zhì)疏松活性篩選

本實(shí)驗(yàn)對(duì)卡氏乳香中分離得到的11個(gè)化合物進(jìn)行了抗骨質(zhì)疏松活性篩選，采用體外核因子-κB配體受體致活劑（receptor activator of nuclear factor kappa-B ligand，RANKL）誘導(dǎo)小鼠骨髓原代巨噬細(xì)胞（bone marrow-derived macrophages，BMMs）形成成熟破骨細(xì)胞模型，進(jìn)行抗骨質(zhì)疏松活性評(píng)估。將細(xì)胞接種于96孔板中，接種12 h，整板更換為新的α-MEM完全培養(yǎng)基，實(shí)驗(yàn)設(shè)對(duì)照組、模型組和給藥組（濃度分別為1、10 μmol/L）。對(duì)照組加入巨噬細(xì)胞集落刺激因子（macrophage colony-stimulating factor，MCSF）的原代小鼠巨噬細(xì)胞，模型組加入誘導(dǎo)因子MCSF以及RANKL的小鼠原代巨噬細(xì)胞，給藥組在模型組的基礎(chǔ)上分別加入1、10 μmol/L化合物溶液。每48小時(shí)換藥1次，直至模型組出現(xiàn)明顯的細(xì)胞融合為止，即得破骨細(xì)胞，棄去培養(yǎng)基，加入500 μL預(yù)冷的固定劑在冰上靜置10 min，加入2 mL PBS 稀釋固定劑，再除去PBS，加入500 μL乙醇-丙酮（50︰50）溶液，在?30～?20 ℃下培養(yǎng)1 min。用雙蒸水洗3次，加入TRAP染色，在37 ℃培養(yǎng)箱內(nèi)反應(yīng)15～45 min至顯色，在顯微鏡下觀察，結(jié)果見(jiàn)圖4。由此可知，化合物1、3、6和10在1 μmol/L濃度時(shí)具有一定的破骨細(xì)胞抑制活性，化合物1在10 μmol/L濃度時(shí)具有良好的破骨細(xì)胞抑制活性。

圖4 化合物1、3、6和10抑制破骨細(xì)胞活性

5 討論

本實(shí)驗(yàn)對(duì)乳香乙醇提取物中二氯甲烷部分進(jìn)行研究，從中分離得到11個(gè)化合物，其中化合物1是新化合物，化合物5為首次從乳香中分離得到，化合物2和10為首次從該屬植物中分離得到。對(duì)11個(gè)化合物進(jìn)行抗骨質(zhì)疏松活性篩選，結(jié)果顯示化合物1、3、6和10在1 μmol/L濃度時(shí)具有一定的破骨細(xì)胞抑制活性，化合物1在10 μmol/L濃度時(shí)具有良好的破骨細(xì)胞抑制活性。本實(shí)驗(yàn)豐富了乳香化學(xué)成分和生物活性研究，闡釋了其發(fā)揮抗骨質(zhì)疏松活性的物質(zhì)基礎(chǔ)，為其進(jìn)一步開(kāi)發(fā)提供了依據(jù)。

利益沖突 所有作者均聲明不存在利益沖突

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A new norditerpenoid from gum resin of

XU Lian-lian, CHEN Zhi-chao, HE Meng-li, YANG Bao, FU Jian-jiang, CHEN Fang-you, LUO Yong-ming

School of Pharmacy, Jiangxi University of Chinese Medicine, Nanchang 330004, China

To study the chemical compositions ofand their anti-osteoporotic activities.The chemical components were separated by silica gel column chromatography, ODS column chromatography and preparative HPLC, and their structures were analyzed by comparison of spectral data and literature.Eleven compounds were isolated from 95% ethanol extract ofand identified as (1,2,8,9)-1,9-epoxy-8-acetoxy-4,5-dioxo-2-cembranene (1),(1,3,7,11,12)-1,12-epoxy-4- methylenecembr-7-ene-3,11-diol (2), boscartin F (3), boscartene K (4), fidmansumbin-13(17)-en-3,16-diene (5), 3α-acetoxyl-7- oxo-tirucalla-8,24-dien-21-oic acid (6), boscartene N (7), 3β-hydroxy-tirucallic acid (8), α-boswellic acid (9), boscarterol O (10), boscarterol F (11).Compound 1 is a new compound, named boscartenoid A, compound 5 is the first time isolated from the species, and compounds 2 and 10 are the fiist time isolated from this genus. Eleven compounds were screened for anti-osteoporotic activity and compounds 1, 3, 6 and 10 were found to have good inhibitory activity against osteoclasts.

Birdw.;terpenoids; norditerpenoids; (1,2,8,9)-1,9-epoxy-8-acetoxy-4,5-dioxo-2-cembranene; boscartenoid A

R284.1

A

0253 - 2670(2023)18 - 5833 - 07

10.7501/j.issn.0253-2670.2023.18.001

2023-05-03

國(guó)家自然科學(xué)基金項(xiàng)目（82060697）；江西中醫(yī)藥大學(xué)校級(jí)科技創(chuàng)新團(tuán)隊(duì)發(fā)展計(jì)劃（CXTD22015）；學(xué)校博士科研啟動(dòng)基金項(xiàng)目（2018BSZR003）

徐蓮蓮，碩士研究生在讀。E-mail: xulianliana@163.com

陳芳有，副教授，從事中藥物質(zhì)基礎(chǔ)研究。Tel: 13037218802 E-mail: tedchenfy@163.com

羅永明，教授，從事中藥物質(zhì)基礎(chǔ)研究。Tel: 13970058758 E-mail: loym999@163.com

[責(zé)任編輯 王文倩]