云 南 植 物 研 究 2008, 30 (6): 725～728 Acta Botanica Yunnanica DOI: 10.3724 SP.J.1143.2008.08160 苏铁蕨的化学成分 方云山 , 杨亚滨 , 杨明惠 , 杨雪琼 , 董刘宏 , 丁中涛 (教育部自然资源药物化学重点实验室, 云南大学化学科学与工程学院, 云南 昆明 650091) 摘要: 从蕨类植物苏铁蕨 ( Brainea insignis) 中分离得到 1个新的苯乙烯基吡喃酮苷, 利用波谱技术鉴定了 其结构。此外, 还得到了1 个已知的苯乙烯基吡喃酮苷和4 个其它已知化合物。同时, 还测定了分离得到 的两个苯乙烯基吡喃酮苷化合物的 DPPH 自由基清除活性。 关键词: 蕨类植物; 苏铁蕨; 苯乙烯基吡喃酮苷; DPPH 自由基清除活性 中图分类号: Q 946 文献标识码: A 文章编号: 0253 -2700 (2008) 06- 725- 04 Chemical Constituents from the Fern Brainea insignis (Blechnaceae) FANG Yun-Shan, YANG Ya-Bing, YANG Ming-Hui, YANG Xue-Qiong, DONG Liu-Hong, DING Zhong-Tao** ( Key Laboratory of Medicinal Chemistry for Natural Resource, Ministry of Education, School of Chemical Science and Technology, Yunnan University, Kunming650091, China) Abstract: A new styryl-2-pyrone glucoside (1) and a known styryl-2-pyrone glucoside (2) , together with four other known compounds (3 -6) , were isolated from the ethanol extract of the fern Brainea insignis .The structures of these compounds were determined by means of spectroscopic analysis including1D and 2D NMR, HR-MS . And the capacities to scavenge against DPPH radical of two styryl-2-pyrone glucosides were assayed . Key words: Fern; Brainea insignis; Styryl-2-pyrone glycoside; DPPH radical scavenging activity Braniea insignis (Hook .) J .Smith is a monotype known styryl-2-pyrone glucoside (2 ), and four other genus plant belonging to Blechnaceae . As an ancient known compounds (3 - 6), from Brainea insignis (Fig . fern species, its early fossils belong to the Mesozoic 1) . And the activities of two styryl-2-pyrone glucosides era, near 400 million years ago . Now it is mainly dis- in scavenging DPPH radical were also reported . tributed in the South of China, and used as a Chinese Compound 1 was obtained as a yellow amorphous traditional medicinal herb for the treatment of common powder . The positive-ion HRESI-MS spectrum gave a cold, suffer burn, trauma bleeding, ascarid disease [M + H]+ ion peak at m z 393.1168 ( calcd . m z etc . (Wu, 1990) .A new flavonoid glycoside and four 393.1186), indicating a molecular formula of C H 19 20 lignans have been isolated from Brainea insignis (Mu- O , with ten degree of unsaturation . In addition, the 9 rakami et al., 1986; Wada et al., 1992) . In the pr- + ESI-MS gave a [M+ H-162] ion at m z 231, which esent paper, we report the isolation and structural elu- showed that compound 1 might be a glycoside . Its 1 H cidation of a new styryl-2-pyrone glucoside (1 ) , a 13 NMR and C NMR spectra (Table 1) confirmed that it Foundation item: The Natural Science Foundation of China and Yunnan Province (30560178) Author for correspondence; Tel: +86-0871-5033726; E-mail: [email protected] Received date: 2008- 08-19, Accepted date: 2008-10-15 作者简介: 方云山 (1980-) 男, 硕士, 主要从事天然产物化学研究, 现在昆明学院工作。 7 2 6 云 南 植 物 研 究 30卷 Fig . 1 Structures of compounds 1-4 Table 1 1H NMR (500 MHz) and 13C NMR (125 MHz) 5′), indicating the presence of a p-disubstituted phe- data of compound 1 in C D N 5 5 nyl ring . And the 1 H NMR signals of a trans double No . δ δ C H bond was noted atδ6.67 (1H, d, J = 16.4 Hz, H - 2 165.1 ww 8′), 7.53 (1H, d, J = 16.4 Hz, H - 7′) . The HM- 3 94.0 6.21 (1H, s) we 4 170.3 BC spectrum showed the correlations (Fig . 2) between ww 5 101.8 6.22 (1H, s) ww H - 8′and C - 1′(δ128.7) suggesting the presence of 6 162.0 ww an E-styryl moiety .The 2-pyrone unit was suggested by 1′ 128.7 iw 2′ 131.5 7.48 (1H, d, 7.8) a detail comparison of13C NMR and 1 H NMR data with iwf 3′ 118.4 7.14 (1H, d, 7.8) iwf the literatures ( Kraut et al., 1996; Mcglacken and 4′ 162.3 iw 5′ 118.4 7.14 (1H, d, 7.8) Fairlamb, 2005), and was identified as a 4, 6-disub- iwf 6′ 131.5 7.48 (1H, d, 7.8) stituted 2-pyrone with two proton signals at δ6.22 (H iwf 7′ 136.8 7.53 (1H, d, 16.4) iw 8′ 118.1 6.67 (1H, d, 16.4) iw 1″ 102.7 5.73 (1H, d, 7.4) iwf 2″ 76.1 4.27 (1H, br d, 6.6) ie 3″ 80.0 4.30 (1H, m) ie 4″ 72.5 4.32 (1H, m) ie 5″ 80.9 4.11 (1H, m) ie 6″ 63.7 4.34 (1H, br d, 11.2) ie 4.29 (1H, br d, 12.2) was a mono-O-β-glucoside with the anomeric proton atδ 5.73 (1H, d, J = 7.4 Hz, H - 1″) and the anomeric carbon atδ102.7 (C - 1″) .The 1 H NMR spectrum ex- hibited an AA′BB′system atδ7.48 (2H, d, J = 7.8 Fig .2 Key HMBC correlations of compound 1 Hz, H - 2′, 6′), 7.14 (2H, d, J= 7.8 Hz, H - 3′, 6期 FANG Yun-Shan et al.: Chemical Constituents from the Fern Brainea insignis (Blechnaceae) 7 27 -5, s) and 6.21 (H - 3, s) in 1 H NMR spectrum . Table 2 Radical DPPH scavenging activity The HMBC correlations clearly revealed that H - 7′in Concentration Scavenging activity (SC% ) Rutin the E-styryl moiety was long-range coupled to the C - 6 (mg ml) Compound1 Compound 2 R 0.01 75.78 6.17 15.52 p in 2-pyrone unit, and the anomeric proton (H - 1″) of 0.03 90.71 17.87 42.95 the glucose moiety was long-range coupled to the C- 4 0.06 92.06 29.07 73.43 (δ171.3) . Therefore, compound 1 was determined to be bisnoryangonin-4-O-β-D- glucopyranoside . Our previous investigation showed that the extract Compound 2 was also obtained as a yellow amor- of B. insignis exhibited high phenolic content and phous powder . It displayed a [M+ H]+ peak at m z strong radical DPPH scavenging activity (Ding et al., 409.1117 (calcd . m z 490.1135) in the positive ion 2008) . In this work, all the substances showed activity HRESIMS, indicating a molecular formula of C H against the radical DPPH, but rutin was more active 19 20 O . Compared with 1, all of the 13 C NMR data were than compound 1 and 2 at the tested contraction . Be- 10 very similar to that of the compound 1, except for the cause compound 2 bears two phenolic hydroxyl groups signalsδ149.2 (C - 4′) , δ146.5 (C - 3′) . This in- on molecular skeleton, its observed activity was stron- dicated that there are two o-hydroxyls in E-styryl moi- ger than that of compound 1 which only has one phe- ety of compound 2 . In 1 H NMR spectrum, the signals nolic hydroxyl group in its structure .This indicted that ofδ6.96 (1H, d, J = 6.2 Hz, H - 6′), 6.78 (1H, the phenolic hydroxyl group is the mainly active group d, J = 6.2 Hz, H - 5′), 7.05 (1H, s, H - 2′) also for the activity against the radical DPPH . proved this conclusion . Therefore, the structure of compound 2 was established as hispidin-4-O-β-D-glu- Experimental copyranoside, which had been isolated from fern Pteris General Experimental Procedures Optical rotations were ensiformis (Chen et al., 2007) . measured with a Jasco DIP-370 digital polarimeter . 1H, 13 C NMR and 2D NMR spectra were measuredon a Bruker DRX-500 Compound 3 was identified as batatasin (III) by analysis of its 1 H, 13 C NMR and MS spectra (Gao et NMR spectrometer (TMS as internal standard, δin ppm, J in Hz) . HRESIMS were obtained on an agilent G3250AA mass al., 2006) . Compound 4 was identified as ponastero- spectrometer . For column chromatography, silica gel (200 - 300 side A by comparison of its physicochemical and NMR mesh, Qingdao Marine Chemical Inc ., China) .Lichroprep RP- data with the reported in the literature (Zhu et al., 18 (40 - 60μm, Merck, Darmstadt, Germany) and Sephadex 2000; Hikino et al., 1969 ) . And β-sitosterol ( 5), LH-20 (25 - 100μm, Amersham Biosciences, Sweden) were daucosterol ( 6) were identified by comparison with used .A shimazu UV-VIS-PC2410 spectrometer was used inDP- their authentic samples . PH radical scavenging activity test . The research into natural products as health pro- Plant Material The whole plant of Brainea insignis tecting factors against oxidative damage is an interesting (Hook .) J . Smith was collected in February 2006, in Yunnan field .DPPH (1, 1-diphenyl-2-picrylhydrazyl) is a sta- province, P . R . China . A voucher specimen (2006D011) was deposited in the school of Chemical Science and Technology of ble free radical that loses its purple color when accepts Yunnan University, and was identified by Prof . Shu-Gang Lu . an electron from an antioxidant compound (Bao et al., Extraction and Isolation The air-dried plant powder of 2004; Marino et al., 2007) . In this paper, the radi- Brainea insignis (1.4kg) was extracted with EtOH for 24h and cal scavenging capacities of the isolates (1 and 2) from filtered .The filtrate was evaporated in vacuo to give a residue . B. insignis were evaluated by investigating them Then the residue (55g) was subjected to column chromatography against the DPPH radical .The substances were assayed on silica gel, and was eluted with CHCl -MeOH (9∶1) to give 3 at three certain concentration of 0.01, 0.03, 0.06 mg four fractions . Fr . III was subjected to Sephadex LH-20 column ml, and their activities were compared with rutin, a chromatography eluting with MeOH to afford compound 1 ( 25 known natural antioxidant compound .The results were mg), 2 (12 mg) , 4 (15mg), 6 (30mg), and were purified by reported in Table 2 . RP-18 column chromatography elutingwith CH OH-H O (9∶1) . 3 2 7 2 8 云 南 植 物 研 究 30卷 Fr . II was subjected to silica gel column chromatography eluting 2006 . Constituents from Pholidota yunnanensis and their inhibitory with EtOAc-MeOH ( 96∶4) to afford compound 3 (10 mg) . effects onnitricoxide production [J] . ChinTrad Herbal Drugs (中草 药), 37: 492—496 Compound5 (50 mg) was from Fr .I . Hikino H, Rihaya T, TakemotoT, 1969 .Ponasteroside A, a glycoside of Bisnoryangonin-4-O-β-D-glucopyranoside ( 1 ) : yellow insect metamorphosin substance from Pteridium aquilinum var. la- amorphous powder . [α]22.5 - 44.13 ( c0.64, MeOH) .1H- and D tiusculum: structure and absolute configuration [ J] . Tetrahedron, 13C-NMR data see table 1 . HR-ESI-MS: m z 393.1168 [M+ 25: 3909—3917 H] + (calcd .for C H O 393.1186) . 19 21 9 Kraut L, Mues R, Speicher A et al., 1996 .Carboxylatedσ-pyrone deri- DPPH Radical Scavenging Assay DPPH radical scaveng- vatives and flavonoids from the liverwort Dumortiera hirsute [ J] . ing activity was examined by the described method (Bao et al., Phytochemistry, 42: 1693—1698 2004; Marino et al., 2007) . In brief, 3.0 ml each sample in Marino SD, Gala F, Borbone N et al., 2007 . Phenolic glycosides from ethanol was added to 2.0 ml DPPH (2.0×10-4 mol L) solution Foeniculum vulgare fruit and evaluation of antioxidative activity [J] . and incubated at room temperature for 30 min . Then the absor- Phytochemistry, 68: 1805—1812 bance of the reaction mixture was determined at 517 nm on a Mcglacken GP, Fairlamb IJS, 2005 .2-pyrone natural products and mim- shimazu UV-VIS-PC 2410 spectrometer . etics: isolation, characterization and biological activity [ J] . Nat Prod Rep, 22: 369—385 References: Murakami T, Wada H, Tanaka N et al., 1986 .Chemical and chemotax- onomical studies of ferns . LXV . A few new flavonoid glycosides Bao ZJ (保志娟), Yang XQ (杨雪琼) , Ding ZT (丁中涛) et al., [J] . Yakugaku Zasshi, 106: 982—988 2004 . 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