陳　玉, 劉　流, 牛艷芬

(中南民族大學(xué) 化學(xué)與材料科學(xué)學(xué)院，武漢 430073)

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靈芝化學(xué)成分的研究

陳玉, 劉流, 牛艷芬

(中南民族大學(xué) 化學(xué)與材料科學(xué)學(xué)院，武漢 430073)

摘要為研究靈芝的化學(xué)成分，利用溶劑提取法，對(duì)正、反相硅膠柱層析法和半制備高效液相色譜法對(duì)其進(jìn)行了分離純化，并根據(jù)理化性質(zhì)和現(xiàn)代波譜技術(shù)進(jìn)行了結(jié)構(gòu)鑒定.結(jié)果表明：從靈芝乙醇提取物分離得到的10個(gè)化合物，分別鑒定為antiquol C (1)、ganoderic acid Y (2)、7-oxo-ganoderic acid Z3 (3)、7-oxo-ganoderic acid Z (4)、ganoderic acid B9 (5)、ganoderic acid C2 (6)、3β,7β-dihydroxy-11,15,23-trioxo-lanost-8,16-dien-26-oic acid (7)、ergosterol (8)、5α,8α-epidioxyergosta-6, 22-dien-3β-ol (9)、6, 9-epidioxy-ergosta-7,22-dien-3β-ol (10)，其中化合物(1)為首次從該屬植物中分離得到.

關(guān)鍵詞靈芝；化學(xué)成分；三萜；甾體

靈芝(Ganodermalucidum)為多孔菌科赤芝或紫芝的干燥子實(shí)體，別名有萬(wàn)年蕈、瑞草、秀草等，是我國(guó)一種傳統(tǒng)的的大型藥用真菌，屬于擔(dān)子菌綱(Basidiomycetes)多孔菌科(Polyporaceae)靈芝亞科(Ganodermatoideae)靈芝屬(Ganoderma)[1].傳統(tǒng)中醫(yī)認(rèn)為靈芝具有補(bǔ)肝益氣、延年益壽的功效.近年來(lái)從靈芝中分離到三萜類化合物已有100種，其中三萜類化合物主要是高度氧化的羊毛甾烷衍生物[2].現(xiàn)代藥理研究表明：靈芝三萜類化合物具有抗腫瘤、抗病毒等作用[3].為了進(jìn)一步深入了解其有效成分，本文對(duì)靈芝的三萜和甾醇成分進(jìn)行了系統(tǒng)的研究.

1材料與方法

1.1儀器與試劑

核磁共振波譜儀(Bruker AM-400型, 布魯克公司)，高效液相色譜儀(Ultimate 3000半制備型, 美國(guó)戴安公司)，VWD，DAD檢測(cè)器，半制備型色譜柱為5C18-MS-II (250 mm×10 mm, 5 μm).

靈芝購(gòu)于云南西雙版納，由云南西雙版納傣族自治州民族醫(yī)藥研究所趙應(yīng)紅主任藥師鑒定為靈芝屬(Ganoderma)靈芝(G.Lucidum).50 μm反相材料ODS(日本YMC公司)；300～400目硅膠和硅膠H-TLC薄層預(yù)制板(青島海洋化工廠)，硫酸香草醛顯色劑(硫酸、香草醛、乙醇、水 分別為8 mL、0.5 g、10 mL、20 mL)，色譜甲醇(美國(guó)Tedia試劑公司)，其余試劑均為分析純，由國(guó)藥集團(tuán)化學(xué)試劑有限公司和天津博迪化工有限公司生產(chǎn).

1.2提取與分離

取靈芝粉碎后稱重1.4 kg，用95%乙醇室溫浸泡3次，每次24 h，合并3次提取液.真空抽濾得濾液，旋干濃縮，得提取物46.7 g.將此靈芝乙醇提取物經(jīng)硅膠柱層析分離，用環(huán)己烷∶丙酮(50∶1→0∶1)梯度洗脫，TLC檢測(cè)后合并相同組分，共收集7個(gè)組分(Fr.A～G).Fr.B經(jīng)過(guò)正、反相硅膠柱色譜分離得到化合物8(200 mg)和9(20 mg).Fr.D經(jīng)過(guò)反復(fù)的正、反相硅膠柱色譜分離得到化合物1(10 mg)、2(22 mg)、3(19 mg)和4(30 mg).Fr.E經(jīng)過(guò)反復(fù)的正相硅膠柱色譜分離得到化合7(6 mg).Fr.F經(jīng)過(guò)反復(fù)的正、反相硅膠柱色譜和半制備HPLC得到化合物5(12 mg)、6(4 mg)、10(9 mg)，其結(jié)構(gòu)式見(jiàn)圖1.

圖1　化合物1～10的結(jié)構(gòu)式Fig.1　Chemical structures of compound 1-10

2結(jié)果

2.1結(jié)構(gòu)鑒定

化合物1：白色粉末，分子式為C30H48O,1H-NMR (400MHz, C5D5N)：δH5.61(1H, d,J=5.6 Hz, H-7), 5.44(1H, d,J=5.6 Hz, H-11), 5.27(1H, t,J=7.0 Hz, H-24), 3.51(1H, dd,J=8.4, 7.1 Hz, H-3), 1.71(3H, s, H-26), 1.66(3H, s, H-27), 1.24(3H, s, H-28), 1.16(3H, s, H-19), 1.12(3H, s, H-29), 1.02(3H, s, H-30),1.00(3H, d,J=5.2 Hz, H-21), 0.71(3H, s, H-18).13C-NMR(100 MHz, C5D5N)數(shù)據(jù)見(jiàn)表1.以上數(shù)據(jù)與文獻(xiàn)報(bào)道基本一致[4]，故鑒定化合物1為antiquol C.

化合物2：白色粉末，分子式C30H46O3,1H-NMR(400MHz, C5D5N)：δH5.61(1H, d,J=4.8 Hz, H-7), 5.44(1H, d,J=5.6 Hz, H-11), 5.23(1H, m, H-24), 3.50(1H, dd,J=8.6, 6.9 Hz, H-3), 2.15(3H, s, H-27), 1.25(3H, s, H-29), 1.17(3H, s, H-19), 1.13(3H, s, H-28), 1.03(3H, s, H-30), 1.00(3H, d,J=6.4Hz, H-21), 0.70(3H, s, H-18).13C-NMR(100 MHz, C5D5N)數(shù)據(jù)見(jiàn)表1.以上數(shù)據(jù)與文獻(xiàn)報(bào)道基本一致[4]，故鑒定化合物2為ganoderic acid Y.

化合物3：白色粉末，分子式為C30H46O5,1H-NMR(400MHz, C5D5N)：δH6.35(1H, m, H-24), 4.98(1H, d,J=5.6 Hz, H-15), 3.57(1H, dd,J=11.6, 4.4 Hz, H-3), 2.12(3H, s, H-27), 1.72(3H, s, H-19), 1.24(3H, s, H-29), 1.19(3H, s, H-18), 1.15(3H, s, H-28), 1.12(3H, s, H-30), 1.01(3H, d,J=5.2 Hz, H-21).13C-NMR(100 MHz, C5D5N)數(shù)據(jù)見(jiàn)表1.以上數(shù)據(jù)與文獻(xiàn)報(bào)道基本一致[5]，故鑒定化合物3為7-oxo-ganoderic acid Z3.

化合物4：白色粉末，分子式為C30H46O4,1H-NMR(400MHz, C5D5N)：δH6.13(1H, m, H-24), 3.47(1H, dd,J=10.8, 6.0 Hz, H-3), 2.14(3H, s, H-27), 1.17(3H, s, H-19), 1.14(3H, s, H-29), 1.12(3H, s, H-28), 1.08(3H, s, H-30), 1.023H, d,J=6.0 Hz, H-21), 0.72(3H, s, H-18).13C-NMR(100 MHz, C5D5N)數(shù)據(jù)見(jiàn)表1.以上數(shù)據(jù)與文獻(xiàn)報(bào)道基本一致[6]，故鑒定化合物4為7-oxo-ganoderic acid Z.

化合物5：白色粉末，分子式為C30H46O7.1H-NMR(400MHz, C5D5N)：δH4.92(1H, dd,J=8.8, 6.4 Hz, H-15), 4.86(1H, br d,J=6.4 Hz, H-7), 3.56(1H, dd,J=11.6, 4.8 Hz, H-3), 3.48(1H, dt,J=13.6,3.6 Hz, H-1), 3.34(1H, m, H-25), 3.15(1H, dd,J=17.6, 8.8 Hz, H-24), 3.01(1H, br d,J=17.6Hz, H-12), 2.64～2.58(1H,m, H-12), 2.64～2.58(1H, m, H-24), 2.39(1H, dd,J= 16.0, 9.6, H-22), 2.17～2.08(2H, m, H-16), 2.201H, m, H-20), 2.15(1H, m, H-6), 2.08～1.93(2H, m, H-2), 2.01(1H, m, H-5), 1.83(1H, m, H-6), 1.78(1H, m, H-17), 1.62(3H, s, H-30), 1.44(1H, m, H-1), 1.37(1H, d,J=7.2Hz, H-27), 1.35(3H, s, H-28), 1.30(3H, s, H-19), 1.14(3H, s, H-29), 0.99(3H, d,J=6.8 Hz, H-21), 0.97(3H, s, H-18).13C-NMR(100 MHz, C5D5N)數(shù)據(jù)見(jiàn)表1.以上數(shù)據(jù)與文獻(xiàn)報(bào)道基本一致[7]，故鑒定化合物5為ganoderic acid B9.

化合物6：白色粉末，分子式為C30H46O7.1H-NMR(400MHz, C5D5N)：δH5.26(1H, br t,J=8.0 Hz, H-15), 4.99(1H, dd,J=10.0, 8.4 Hz, H-7), 3.54(1H, dd,J=11.2, 4.4 Hz, H-3), 3.31(1H, m, H-25), 3.13(1H, m, H-1), 3.09(1H, m, H-24), 3.02(1H, br d,J=15.2Hz, H-12), 2.70(1H,J=15.2Hz, H-12), 2.57(1H, m, H-22), 2.56(1H, m, H-24), 2.41(1H, m, H-6), 2.31(1H, m, H-22), 2.18(1H, m, H-20), 2.14～1.94(2H, m, H-16), 1.98(1H, m, H-6), 1.92(2H, m, H-2), 1.91(1H, m, H-17), 1.55(3H, s, H-18), 1.53(3H, s, H-28), 1.37(1H, d,J=6.8Hz, H-27), 1.29(3H, s, H-30), 1.19(1H, m, H-5), 1.15(1H, m, H-1), 1.12(3H, s, H-29), 0.98(3H, d,J=6.0 Hz, H-21).13C-NMR(100 MHz, C5D5N)數(shù)據(jù)見(jiàn)表1.以上數(shù)據(jù)與文獻(xiàn)報(bào)道基本一致[8]，故鑒定化合物6為ganoderic acid C2.

化合物7：白色粉末，分子式C30H42O7.1H-NMR(400MHz, C5D5N)：δH5.98(1H, s, H-16), 5.14(1H, dd,J=9.6, 8.4 Hz, H-7), 3.53(1H, dd,J=7.2, 4.8 Hz, H-3), 3.26(1H, m, H-1), 3.13(1H, m, H-12), 3.12(1H, m, H-24), 3.10(1H, m, H-20), 3.09(3H, d,J=7.2 Hz, H-27), 2.95(1H, dd,J=17.7, 4.4 Hz, H-22), 2.82(1H, dd,J=17.2, 6.4 Hz, H-22), 2.73(1H,J=16.0 Hz, H-12), 2.69(1H, m, H-25), 2.56(1H, dd,J=17.2, 4.4 Hz, H-24), 2.36(1H, dd,J=13.2, 7.6 Hz, H-6), 1.92(2H, m, H-2), 1.87(1H, m, H-6), 1.48(3H, s, H-18), 1.41(3H, s, H-19), 1.29(3H, s, H-28), 1.25(1H, m, H-1), 1.18(1H, m, H-5), 1.11(3H, s, H-29), 1.10(3H, d,J=7.2 Hz, H-21).13C-NMR(100 MHz, C5D5N)數(shù)據(jù)見(jiàn)表1.以上數(shù)據(jù)與文獻(xiàn)報(bào)道基本一致[9]，故鑒定化合物7為3β,7β-dihydroxy-11,15,23-trioxo-lanost-8,16-dien-26-oic acid.

化合物8：白色粉末，分子式C28H44O,1H-NMR(400MHz, C5D5N)：δH5.73(1H, dd,J=6.4, 2.0 Hz, H-6), 5.52(1H, dd,J=5.6, 2.8 Hz, H-7), 5.29(1H, m, H-23), 5.25(1H, m, H-22), 3.97(1H, m, H-3), 1.11(3H, d,J=6.4 Hz, H-21), 1.06(3H, s, H-19), 1.00(3H, d,J=7.2 Hz, H-28), 0.90(3H, d,J=6.8 Hz, H-27), 0.89(3H, d,J=6.8 Hz, H-26), 0.69(3H, s, H-18).13C-NMR(100 MHz, C5D5N)數(shù)據(jù)見(jiàn)表1.以上數(shù)據(jù)與文獻(xiàn)報(bào)道基本一致[10]，故鑒定化合物8為ergosterol.

化合物9：針狀結(jié)晶(甲醇)，分子式為C28H46O3,1H-NMR(400MHz, C5D5N)：δH6.55(1H, d,J=8.4 Hz, H-6), 6.35(1H, d,J=8.4 Hz, H-7), 5.31(1H, m, H-23), 5.22(1H, m, H-22), 4.43(1H, m, H-3), 1.05(3H, d,J=6.4 Hz, H-21), 0.98(3H, d,J=6.8 Hz, H-28), 0.91(3H, s, H-19), 0.89(3H, d,J=6.8 Hz, H-27), 0.88(3H, d,J=6.8 Hz, H-26), 0.79(3H, s, H-18).13C-NMR(100 MHz, C5D5N)數(shù)據(jù)見(jiàn)表1.以上數(shù)據(jù)與文獻(xiàn)報(bào)道基本一致[11]，故鑒定化合物9為5α,8α-epidioxyergosta-6,22-dien-3β-ol.

化合物10：白色粉末，分子式為C28H44O2,1H-NMR(400MHz, C5D5N)：δH6.36(1H, d,J=5.6 Hz, H-7), 6.10(1H, d,J=4.8 Hz, H-23), 5.77(1H, m, H-22), 4.87(1H, m, H-6), 4.35(1H, m, H-3), 1.56(3H, s, H-19), 1.08(3H, d,J=6.8 Hz, H-21), 0.97(3H, d,J=6.8 Hz, H-28), 0.88(3H, d,J=6.4 Hz, H-27), 0.87(3H, d,J=6.8 Hz, H-26), 0.67(3H, s, H-18).13C-NMR(100 MHz, C5D5N)數(shù)據(jù)見(jiàn)表1.以上數(shù)據(jù)與文獻(xiàn)報(bào)道基本一致[12]，故鑒定化合物10為6,9-epidioxy-ergosta-7,22-dien-3β-ol.

表1　化合物1～10的13C-NMR數(shù)據(jù)

3結(jié)語(yǔ)

通過(guò)對(duì)靈芝的系統(tǒng)分離，得到10種化合物，分別鑒定為antiquol C(1)、ganoderic acid Y(2)、7-oxo-ganoderic acid Z3(3)、7-oxo-ganoderic acid Z(4)、ganoderic acid B9(5)、ganoderic acid C2(6)、3β,7β-dihydroxy-11,15,23-trioxo-lanost-8,16-dien-26-oic acid(7)、ergosterol(8)、5α, 8α-epidioxyergosta-6, 22-dien-3β-ol(9)、6, 9-epidioxy-ergosta-7,22-dien-3β-ol(10)，其中化合物1為首次從該屬植物中分離得到，為進(jìn)一步研究和開(kāi)發(fā)該藥物提供了實(shí)驗(yàn)基礎(chǔ)和理論依據(jù).

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Chemical Constituents ofGanodermalucidum

ChenYu,LiuLiu，NiuYanfen

(College of Chemistry and Materials Science，South-Central University for Nationalities， Wuhan 430074，China)

AbstractTo study the chemical constituents ofGanodermalucidum, the constituents were isolated and purified by the silica gel column chromatography, ODS and semi-preparative HPLC.Their structures were elucidated on the basis of chemical and spectroscopic analysis. Ten compounds were isolated fromG.lucidumand identified as: antiquol C (1), ganoderic acid Y (2), 7-oxo-ganoderic acid Z3 (3), 7-oxo-ganoderic acid Z (4), ganoderic acid B9 (5), ganoderic acid C2 (6), 3β,7β-dihydroxy-11,15,23-trioxo-lanost-8,16-dien-26-oic acid (7), ergosterol (8), 5α,8α-epidioxyergosta-6, 22-dien-3β-ol (9), 6,9-epidioxy-ergosta-7,22-dien-3β-ol (10), among them compound 1 was isolated from the genusGanodermafor the first time.

KeywordsGanodermalucidum; chemical constituents; triterpenoids; steroids

收稿日期2015-09-23

作者簡(jiǎn)介陳玉(1970-),女,副教授,博士,研究方向:天然藥物化學(xué), E-mail: chenyuwh888@126.com

基金項(xiàng)目國(guó)家自然科學(xué)基金資助項(xiàng)目(31370379)

中圖分類號(hào)R914.4

文獻(xiàn)標(biāo)識(shí)碼A

文章編號(hào)1672-4321(2016)02-0010-05