Article(id=1190373736432894697, tenantId=1146029695717560320, journalId=1189982191388893191, issueId=1190332325088039709, articleNumber=null, orderNo=null, doi=10.16438/j.0513-4870.2024-1190, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=research-article, receivedDate=1733155200000, receivedDateStr=2024-12-03, revisedDate=1739462400000, revisedDateStr=2025-02-14, acceptedDate=null, acceptedDateStr=null, onlineDate=1761736814840, onlineDateStr=2025-10-29, pubDate=1746979200000, pubDateStr=2025-05-12, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1761736814840, onlineIssueDateStr=2025-10-29, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1761736814840, creator=13701087609, updateTime=1761736814840, updator=13701087609, issue=Issue{id=1190332325088039709, tenantId=1146029695717560320, journalId=1189982191388893191, year='2025', volume='60', issue='5', pageStart='1183', pageEnd='1572', issueExtLink='null', onlineDate='null', pubDate='null', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=1, specialIssue=null, createTime=1761726941606, creator=13701087609, updateTime=1761813457266, updator=13701087609, preIssue=null, nextIssue=null, ext={EN=IssueExt(id=1190695198163354009, tenantId=1146029695717560320, journalId=1189982191388893191, issueId=1190332325088039709, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1190695198163354010, tenantId=1146029695717560320, journalId=1189982191388893191, issueId=1190332325088039709, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null}, startPage=1485, endPage=1489, ext={EN=ArticleExt(id=1190373736709718762, articleId=1190373736432894697, tenantId=1146029695717560320, journalId=1189982191388893191, language=EN, title=A new sesquiterpenes from Pteris wallichiana and its anticancer activity, columnId=1190335348761793317, journalTitle=Acta Pharmaceutica Sinica, columnName=Original Articles, runingTitle=null, highlight=null, articleAbstract=

The methanol extract of Pteris wallichiana was separated and purified by MCI gel, sephadex LH 20, flash C18 and silica gel column chromatography combined with semi-pre HPLC. The chemical structures of the isolated compounds were identified by MS, IR, NMR, etc. Five sesquiterpene compounds were isolated from Pteris wallichiana and identified as 6,7-tetrahydrofuran-(2S, 3S)-pterosin C-3-O-β-D-(6′-acetyl)-Glu (1), (2S, 3S)-pterosin C-3-O-β-D-Glu (2), (2S)-pterosin A (3) and (2S)-13-hydroxyl-pterosin A (4), (2R, 3S)-2-hydroxyl-pterosin C (5). Compound 1 is a new sesquiterpene, compounds 3-5 were isolated for the first time. In vitro bioactivity assay showed that compound 1 was able to inhibit the proliferation of 4T1 and EMT6 cells, and possessed significant anti-triple-negative breast cancer bioactivity.

, correspAuthors=Duo CAO, authorNote=null, correspAuthorsNote=null, copyrightStatement=Copyright ©2025 Acta Pharmaceutica Sinica. All rights reserved., copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=null, pdfFileSize=null, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=null, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=null, mapNumber=null, authorCompany=null, fund=null, authors=null, authorsList=Shi-jie REN, Hong-ke WEI, Xin-xin CHENG, Jia-qi WANG, Xiao-ting QIAO, Xiao-min WANG, Zi-yu LÜ, Duo CAO), CN=ArticleExt(id=1190373851180663563, articleId=1190373736432894697, tenantId=1146029695717560320, journalId=1189982191388893191, language=CN, title=西南凤尾蕨中一个新的倍半萜及其抗癌活性研究, columnId=1190335348896011050, journalTitle=药学学报, columnName=研究论文, runingTitle=null, highlight=null, articleAbstract=

采用MCI树脂、葡聚糖凝胶Sephadex LH 20、Flash C18制备色谱和半制备液相制备色谱等方法, 对西南凤尾蕨根部甲醇提取物进行较为系统的分离纯化, 通过质谱、红外光谱和核磁共振等方法, 对分离到的化合物进行结构鉴定。从西南凤尾蕨中分离鉴定了5个倍半萜类化合物, 分别为6,7-四氢吡喃环-(2S, 3S)-蕨素C-3-O-β-D-(6′-acetyl)-葡萄糖苷(1)、(2S, 3S)-蕨素C-3-O-β-D葡萄糖苷(2)、(2S)-蕨素A (3)、(2S)-13-羟基蕨素A (4) 和(2R, 3S)-2-羟基蕨素C (5)。其中, 化合物1为新的三环倍半萜, 化合物3~5首次从西南凤尾蕨中分离。体外生物活性实验表明, 化合物1能够抑制4T1和EMT6细胞增殖, 具有显著抗三阴性乳腺癌生物活性。

, correspAuthors=曹朵, authorNote=null, correspAuthorsNote=
*曹朵,E-mail:
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#共同第一作者.

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Medical School of Yan'an University, Yan′an 716000, China), AuthorCompanyExt(id=1190694361676526071, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1190373736432894697, companyId=1190694361651360245, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=2.延安大学医学院, 陕西 延安 716000)])], figs=[ArticleFig(id=1190694371944182340, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1190373736432894697, language=EN, label=null, caption=null, figureFileSmall=jL/UydUqkGbC2P67ofqdyw==, figureFileBig=VhvBC/yZRi5gI6v1Z8KGng==, tableContent=null), ArticleFig(id=1190694372103565894, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1190373736432894697, language=CN, label=Figure 1, caption= Structure of compound <strong>1</strong> and its key <sup>1</sup>H-<sup>1</sup>H COSY, HMBC correlations , figureFileSmall=jL/UydUqkGbC2P67ofqdyw==, figureFileBig=VhvBC/yZRi5gI6v1Z8KGng==, tableContent=null), ArticleFig(id=1190694372313281096, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1190373736432894697, language=EN, label=null, caption=null, figureFileSmall=null, figureFileBig=null, tableContent=
No. δC δH (J in Hz) HMBC
1 205.7 - H-2
2 52.0 2.64 (1H, d, J = 3.8 Hz) H-3, C-1, 3, 11
3 83.1 4.73 (1H, d, J = 3.8 Hz) H-2, C-2, 9
4 126.2 7.50 (1H, s) -
5 144.7 - H-12
6 136.4 - -
7 139.6 - H-15
8 131.4 - -
9 151.4 - H-3
10 - - -
11 13.8 1.26 (3H, d, J = 7.8 Hz) C-2, H-2
12 21.2 2.44 (3H, s) C-5
13 30.1 3.02 (2H, t, J = 8.0 Hz) C-6, 14, H-14
14 63.1 4.31 (2H, s) C-13, H-13
15 65.7 3.16 (2H, s) C-7
1′ 106.4 4.50 (1H, d, J = 8.0 Hz) C-3, H-2′, C-2′
2′ 74.1 3.01 (1H, m) C-3′
3′ 77.2 3.20 (1H, m) C-2′, 4′
4′ 70.5 3.09 (1H, m) C-3′, 5′
5′ 77.4 3.21 (1H, m) C-4′, 6′
6′ 63.1 3.49 (1H, dt, J = 12.2, 6.0 Hz), 3.72 (1H, ddd, J = 12.2, 5.2, 2.2 Hz) C-5′, 1″
1″ 170.8 - C-6′, 2″, H-2″
2″ 21.2 2.0 (3H, s) C-1″
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1H NMR (800 MHz) and 13C NMR (200 MHz) data of compound 1 in DMSO-d6

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No. δC δH (J in Hz) HMBC
1 205.7 - H-2
2 52.0 2.64 (1H, d, J = 3.8 Hz) H-3, C-1, 3, 11
3 83.1 4.73 (1H, d, J = 3.8 Hz) H-2, C-2, 9
4 126.2 7.50 (1H, s) -
5 144.7 - H-12
6 136.4 - -
7 139.6 - H-15
8 131.4 - -
9 151.4 - H-3
10 - - -
11 13.8 1.26 (3H, d, J = 7.8 Hz) C-2, H-2
12 21.2 2.44 (3H, s) C-5
13 30.1 3.02 (2H, t, J = 8.0 Hz) C-6, 14, H-14
14 63.1 4.31 (2H, s) C-13, H-13
15 65.7 3.16 (2H, s) C-7
1′ 106.4 4.50 (1H, d, J = 8.0 Hz) C-3, H-2′, C-2′
2′ 74.1 3.01 (1H, m) C-3′
3′ 77.2 3.20 (1H, m) C-2′, 4′
4′ 70.5 3.09 (1H, m) C-3′, 5′
5′ 77.4 3.21 (1H, m) C-4′, 6′
6′ 63.1 3.49 (1H, dt, J = 12.2, 6.0 Hz), 3.72 (1H, ddd, J = 12.2, 5.2, 2.2 Hz) C-5′, 1″
1″ 170.8 - C-6′, 2″, H-2″
2″ 21.2 2.0 (3H, s) C-1″
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Compound IC50/µmol·L-1
4T1 EMT6 U87 U251 TE10 ECA109
1 8.61 8.79 18.34 21.01 > 50 > 50
2 33.65 29.54 > 50 37.78 > 50 > 50
3 > 50 > 50 > 50 > 50 > 50 > 50
4 > 50 > 50 > 50 > 50 > 50 > 50
5 > 50 > 50 39.01 36.98 > 50 > 50
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Screening results for antitumor activity of pterosins 1-5

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Compound IC50/µmol·L-1
4T1 EMT6 U87 U251 TE10 ECA109
1 8.61 8.79 18.34 21.01 > 50 > 50
2 33.65 29.54 > 50 37.78 > 50 > 50
3 > 50 > 50 > 50 > 50 > 50 > 50
4 > 50 > 50 > 50 > 50 > 50 > 50
5 > 50 > 50 39.01 36.98 > 50 > 50
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西南凤尾蕨中一个新的倍半萜及其抗癌活性研究
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任世杰 1, # , 魏宏珂 1, # , 成欣欣 1 , 王嘉琪 2 , 乔晓婷 2 , 王晓敏 2 , 吕子玉 1 , 曹朵 1, *
药学学报 | 研究论文 2025,60(5): 1485-1489
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药学学报 | 研究论文 2025, 60(5): 1485-1489
西南凤尾蕨中一个新的倍半萜及其抗癌活性研究
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任世杰1, #, 魏宏珂1, #, 成欣欣1, 王嘉琪2, 乔晓婷2, 王晓敏2, 吕子玉1, 曹朵1, *
作者信息
  • 1.延安大学生命科学学院, 陕西省黄土高原资源植物研究与利用省市共建重点实验室, 陕西 延安 716000
  • 2.延安大学医学院, 陕西 延安 716000

通讯作者:

*曹朵,E-mail:
A new sesquiterpenes from Pteris wallichiana and its anticancer activity
Shi-jie REN1, Hong-ke WEI1, Xin-xin CHENG1, Jia-qi WANG2, Xiao-ting QIAO2, Xiao-min WANG2, Zi-yu LÜ1, Duo CAO1, *
Affiliations
  • 1. Shaanxi Key Laboratory of Research and Utilization of Resource Plants on the Loess Plateau, College of Life Sciences, Yan′an University, Yan′an 716000, China
  • 2. Medical School of Yan'an University, Yan′an 716000, China
出版时间: 2025-05-12 doi: 10.16438/j.0513-4870.2024-1190
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采用MCI树脂、葡聚糖凝胶Sephadex LH 20、Flash C18制备色谱和半制备液相制备色谱等方法, 对西南凤尾蕨根部甲醇提取物进行较为系统的分离纯化, 通过质谱、红外光谱和核磁共振等方法, 对分离到的化合物进行结构鉴定。从西南凤尾蕨中分离鉴定了5个倍半萜类化合物, 分别为6,7-四氢吡喃环-(2S, 3S)-蕨素C-3-O-β-D-(6′-acetyl)-葡萄糖苷(1)、(2S, 3S)-蕨素C-3-O-β-D葡萄糖苷(2)、(2S)-蕨素A (3)、(2S)-13-羟基蕨素A (4) 和(2R, 3S)-2-羟基蕨素C (5)。其中, 化合物1为新的三环倍半萜, 化合物3~5首次从西南凤尾蕨中分离。体外生物活性实验表明, 化合物1能够抑制4T1和EMT6细胞增殖, 具有显著抗三阴性乳腺癌生物活性。

西南凤尾蕨  /  化学成分  /  倍半萜  /  活性氧  /  三阴性乳腺癌

The methanol extract of Pteris wallichiana was separated and purified by MCI gel, sephadex LH 20, flash C18 and silica gel column chromatography combined with semi-pre HPLC. The chemical structures of the isolated compounds were identified by MS, IR, NMR, etc. Five sesquiterpene compounds were isolated from Pteris wallichiana and identified as 6,7-tetrahydrofuran-(2S, 3S)-pterosin C-3-O-β-D-(6′-acetyl)-Glu (1), (2S, 3S)-pterosin C-3-O-β-D-Glu (2), (2S)-pterosin A (3) and (2S)-13-hydroxyl-pterosin A (4), (2R, 3S)-2-hydroxyl-pterosin C (5). Compound 1 is a new sesquiterpene, compounds 3-5 were isolated for the first time. In vitro bioactivity assay showed that compound 1 was able to inhibit the proliferation of 4T1 and EMT6 cells, and possessed significant anti-triple-negative breast cancer bioactivity.

Pteris wallichiana  /  chemical composition  /  sesquiterpene  /  reactive oxygen species  /  triple negative breast cancer
任世杰, 魏宏珂, 成欣欣, 王嘉琪, 乔晓婷, 王晓敏, 吕子玉, 曹朵. 西南凤尾蕨中一个新的倍半萜及其抗癌活性研究. 药学学报, 2025 , 60 (5) : 1485 -1489 . DOI: 10.16438/j.0513-4870.2024-1190
Shi-jie REN, Hong-ke WEI, Xin-xin CHENG, Jia-qi WANG, Xiao-ting QIAO, Xiao-min WANG, Zi-yu LÜ, Duo CAO. A new sesquiterpenes from Pteris wallichiana and its anticancer activity[J]. Acta Pharmaceutica Sinica, 2025 , 60 (5) : 1485 -1489 . DOI: 10.16438/j.0513-4870.2024-1190
西南凤尾蕨(Pteris wallichana J. Agardh) 为凤尾蕨科(Pteridaceae)、凤尾蕨属(Pteris) 多年生三叉系中大型蕨类植物。生长于海拔800~2 400米林下沟谷, 在中国台湾、海南、云南、四川、西藏等地分布, 在日本、菲律宾、印度等国家也有分布[1]。其性凉、微苦、味涩。常以全草、根茎、叶入药, 具有清热止血、解毒、消肿等功效[2, 3]。《滇药录》中记载: “其叶治小儿高热惊风, 根治疗肝气郁结, 腹部包块, 产后少腹痛, 恶露不尽, 胃脘胀满”[4]。现代研究表明, 西南凤尾蕨含有黄酮类、萜类化合物、甾醇类等物质, 具有降血糖、降血脂、抗炎、免疫调节等生物活性[5-8]。前期预实验发现西南凤尾蕨甲醇提取物具有显著抑制乳腺癌细胞增殖的作用。三阴性乳腺癌作为乳腺癌的一种, 其患病率为10%~30%, 死亡率较高, 临床治疗以化疗为主, 现有药物效果均不理想且存在严重耐药现象[9, 10]。本课题在活性追踪下, 对西南凤尾蕨根部甲醇提取物进行分离、纯化和结构鉴定, 并通过体外细胞实验评价单体物质对癌细胞增殖的抑制作用。
化合物1: 白色粉末, 熔点为215.1 ℃, 可溶于甲醇、氯仿和丙酮等有机溶剂。根据1H NMR和13C NMR以及高分辨HRESI-MS m/z 459.261 3 [M+Na]+ (calcd. for C22H28O9, 459.260 4), 可知其分子式为C22H28O9, 不饱和度为9。从IR光谱中可以看到羰基碳(1 700 cm-1) 以及苯环(1 630, 1 465 cm-1) 特征峰。1H NMR谱显示在δH 2.44 (3H, s, Me-11) 处有一个甲基质子单峰信号, δH 1.26 (3H, d, J = 7.80 Hz) 为一个甲基质子信号双峰信号(Me-10), 在δH 3.16 (2H, s, H-15) 处有一个亚基单峰信号, 还可观察到端基质子δH 4.50 (1H, d, J = 8.0 Hz) 与δC 106.4、77.4、74.1相关, 表明化合物1结构中存在β-吡喃葡萄糖单元, 在HMBC谱中可观察到端基质子与δC 83.1 (C-3) 相关、δH 4.73 (1H, d, J = 3.68 Hz, H-3) 处质子与δC 106.4 (C-1′) 相关、δH 2.0 (3H, s, H-1″) 和δH 3.49 (1H, dt, J = 12.2, 6.0 Hz, H-6′) 均与羰基碳δC 170.8相关, 表明化合物1的C-3位连接6′-acetyl-β-吡喃葡萄糖单元。此外, 13C NMR和DEPT谱还显示14个碳原子核磁共振信号, 其中包括1个羰基信号峰(δC 205.7)、五取代苯环信号峰[δC 126.2 (C-4), 144.7 (C-5), 136.4 (C-6), 139.6 (C-7), 131.4 (C-8), 151.4 (C-9)]、1个sp3次甲基信号峰(δC 52.0, C-2), 4个sp3亚甲基信号峰(δC 32.2, 63.1, 62.5和65.7) 和2个甲基信号峰(δC 13.8和21.2)。紫外吸收在215、260和300 nm处, 红外吸收在1 700、1 600 cm-1处显示了蕨素类化合物的特有吸收峰[11], 通过对化合物1的核磁共振图谱解析, 紫外和红外吸收和文献对比, 推测化合物1的基本骨架是含14或15个碳原子的倍半萜类化合物。同时化合物11H NMR和13C NMR谱数据与pterosine C-3-O-β-葡萄糖苷相似[12], 表明化合物1也是1-茚满酮葡萄糖苷。然而, 与pterosine C-3-O-β-葡萄糖苷不同的是, 化合物113C NMR谱中多1个羰基碳信号峰、1个甲基和1个亚甲基信号峰, 化学位移分别为δC 170.8, 21.2和65.7, 少一个甲基信号峰(δC 24.1), 在HSQC谱中显示δH 3.16 (2H, s, H-15) 与δC 65.7相关、δH 2.0 (3H, s) 与δC 21.2相关, HMBC谱中显示δH 3.16 (2H, s, H-15) 与δC 139.6 (C-7) 相关, 结合分子量分析, C-7、C-8和C-15位化学位移值向低场移动以及不饱和度计算, 推测化合物1的C-6位与C-7位取代基形成一个含氧六元环。对化合物1进行酸水解产生的糖元经气相色谱分析, 证实为D型葡萄糖。1H-1H COSY谱中可观察到δH 2.64 (1H, d, J = 3.8 Hz) 与δH 4.67 (1H, d, J = 3.8 Hz) 相关, 在NOESY谱中不能观察到二者相关, 表明H-2与H-3在异侧, 检测化合物1的母核的比旋光度为$ {\alpha }_{\mathrm{D}}^{20} $ +52 (c 0.5, MeOH), 与(2S, 3S)-pterosine C一致[13]。化合物1的核磁共振数据如表 1所示, 结构如图 1所示。经Scifinder数据库检索, 化合物1为一个新的三元环1H-茚满酮倍半萜类化合物, 命名为6,7-四氢吡喃环-(2S, 3S)-pterosin C-3-O-β-D-(6′-acetyl)-葡萄糖苷。
体外细胞实验检测化合物1~5对EMT6、4T1、U87、U251、TE10和ECA109细胞株增殖的影响, 结果显示化合物1能够显著诱导三阴性乳腺癌细胞EMT6和4T1的增殖, IC50值分别为8.79和8.61 µmol·L-1 (表 2)。
本研究在前期研究基础上, 综合应用分离纯化方法对西南凤尾蕨根部甲醇提取物中化学成分进行较为系统的挖掘, 从中分离鉴定了5个倍半萜类化合物, 其中化合物1为新的三环1-茚满酮倍半萜, 化合物3~5首次从西南凤尾蕨中分离得到。基于前期研究和文献报道, 本研究对分离得到的化合物进行了体外抗癌生物活性评价, 结果表明化合物12对U87、U251、EMT6、4T1细胞的增殖具有显著抑制作用。此外, 化合物1能够诱导EMT6、4T1细胞凋亡(IC50 < 10 µmol·L-1)。总之, 本研究的开展, 不仅丰富了西南凤尾蕨化学成分, 还为后续1-茚满酮倍半萜类化合物的抗癌生物活性研究奠定了基础。
西南凤尾蕨(Pteris wallichiana) 采于云南大理, 经西北大学郭斌教授鉴定为西南凤尾蕨全草, 样本编号2023-23-7, 保存于延安大学生命科学学院特立楼317室。
实验所用到的主要仪器设备有: Agilent Series 1100 SL液质联用仪高分辨ESI, 美国Agilent公司; Bruker Avance 500和AV800型1D和2D核磁共振仪, 德国Bruker公司; Teledyne Isco Combiflash Rf200 Flash分离纯化制备色谱仪器, 皕赫科学仪器上海有限公司; NP7000和NU3000半制备液相色谱仪, 江苏汉邦科技股份有限公司; C18色谱柱(250 mm × 4.6 mm, 5 μm), 日本YMC Co., Ltd.公司; SpectraMax Plus384光吸收酶标仪, 上海美谷分子仪器有限公司; 奥林巴斯IX51荧光倒置显微镜, 基恩士(中国) 有限公司; HF90二氧化碳培养箱, 上海力申科学仪器有限公司; ZF-7紫外分析仪, 上海嘉鹏科技有限公司; DHG-9070A电热恒温鼓风干燥箱, 上海齐欣科学仪器有限公司; 玻璃层析柱(100 mm × 600 mm、80 mm × 500 mm、60 mm × 600 mm、30 mm × 600 mm), 江苏盐城新明特玻璃仪器有限公司; 硅胶(100~200、200~300、300~400目正相硅胶填料) 和GF254薄层硅胶分析板(10 cm × 10 cm), 青岛海湾精细化工有限公司; 葡聚糖凝胶Sephadex LH-20 (500 g, 分离范围100~4 000), 上海源叶生物科技有限公司; Flash C18反相硅胶(40~60 μm, 1 000 g), 上海技舟化工科技有限公司; MCI树脂(型号MCI Gel-CHP 20-P, 粒径75~150 μm), 日本三菱公司; 乙醇、甲醇、石油醚、乙酸乙酯、二氯甲烷、二甲基亚砜(分析纯, 500 mL), 天津市致远化学试剂有限公司。
西南凤尾蕨干燥根部30 kg, 甲醇加热回流提取3次, 回收溶剂后合并滤液得甲醇提取浸膏9 kg, 纯净水分散浸膏后依次用石油醚、二氯甲烷和乙酸乙酯萃取, 得石油醚萃取部位部位340 g、二氯甲烷萃取部位890 g、乙酸乙酯萃取部位2 kg。其中, 乙酸乙酯萃取部位用甲醇溶解后, 按照重量比1∶1.5加100~200目硅胶进行拌样, 通过正相硅胶柱层析, 洗脱液为二氯甲烷∶甲醇= 100∶0~0∶100, 薄层板(TLC) 检测合并相同组分, 获得8个组分Fr. EA-1~8。对Fr. EA-3 (310 g) 进行硅胶柱层析(200~300目硅胶, 二氯甲烷∶乙酸乙酯= 15∶1~1∶1) 薄层色谱检测合并后得7个组分(Fr. EA-3-1~Fr. EA-3-7), Fr. EA-5 (550 g) 也进行硅胶柱层析(200~300目硅胶, 二氯甲烷∶甲醇= 30∶1~5∶1), TLC检测合并得6个组分(Fr. EA-5-1~Fr. EA-5-6)。
Fr. EA-3-3 (102 g) 经MCI树脂柱层析(甲醇-水= 30%~100%), 得Fr. EA-3-3-1~Fr. EA-3-3-6, Fr. EA-3-3-4 (33.5 g) 经Flash C18柱层析, 30%~100%甲醇洗脱, 得Fr. EA-3-3-4-1~Fr. EA-3-3-4-6。其中Fr. EA-3-3-4-2 (60.3 mg) 再经半制备液相色谱(pre-HPLC) 制备, 85.0%甲醇等度洗脱, 得化合物1 (8.3 mg, tR = 18 min) 和化合物2 (14.6 mg, tR = 23 min)。
Fr. EA-5-4 (230.8 g) 经葡聚糖凝胶Sephadex LH 20 (甲醇洗脱) 柱色谱, 得Fr. EA-5-4-1~Fr. EA-5-4-5, Fr. EA-5-4-3 (66.3 g) 经Flash C18柱色谱, 30%~100%甲醇洗脱, 得Fr. EA-5-4-3-1~Fr. EA-5-4-3-5, 其中Fr. EA-5-4-3-3 (74.2 mg) 再经pre-HPLC制备, 93.0%甲醇等度洗脱, 得化合物3 (12.4 mg, tR = 17 min)、化合物4 (11.6 mg, tR = 27 min)、化合物5 (7.9 mg, tR = 24 min)。
化合物1为白色粉末(MeOH), 熔点为215.1 ℃, 可溶于甲醇、氯仿和丙酮等溶剂, 分子式为C22H28O9, $ {\alpha }_{\mathrm{D}}^{20} $ +52 (c 0.5, MeOH); UV (CH3OH) λmax$ $(log ε): 220 (4.0), 在215、260和300 nm处有最大吸收, 为蕨素倍半萜类物质的紫外特征吸收; IR νmax 1 700、1 630、1 465 cm-1, 提示分子中含有羰基和苯环; 1H NMR (800 MHz, DMSO-d6) 和13C NMR (200 MHz, DMSO-d6) 数据见表 1
对化合物1中糖组分的水解和检测按照文献[14]中记载的方法进行, 即将化合物1 (2.0 mg) 在3.0 mL的盐酸二氧六环(1∶1, V/V) 中加热回流2 h, 旋转蒸发仪回收溶剂后用100 µL无水吡啶溶解, 再加200 µL (0.1 mol·L-1) 的半胱氨酸甲酯盐酸盐, 混合物在60 ℃下加热1 h, 再加入3.0 mL的三甲基硅化试剂六甲基二硅氮烷-三甲基氯硅烷-吡啶(HMDA-TMCS, 2∶1∶10, V/V), 在60 ℃条件下继续加热30 min, 反应完全后, 混合物用2 mL环己烷萃取, 萃取物进行GC-MS分析以鉴定糖的构型。对照品单糖(β-D-葡萄糖、β-L-葡萄糖、α-D-葡萄糖、α-L-葡萄糖, 纯度均 > 95%) 购买自百灵威, 对照品也采用上述方法处理后用环己烷萃取, 进行GC-MS分析, 测试结果作为对照。
EMT6、4T1、U87、U251、TE10和ECA109细胞株均来自中国人民解放军空军军医大学基础医学院捐赠。
细胞培养    将细胞在含有10%胎牛血清、1%青-链霉素的DMEM/1640完全培养基中, 在37 ℃、5% CO2培养条件下培养。待细胞长满培养皿底部80%时进行传代和处理。将长满培养皿底的细胞培养基弃去, 后用PBS缓冲液冲洗一次, 加入含EDTA、酚红的胰蛋白酶1~2 mL进行消化约2~3 min, 在显微镜下可以看到贴壁细胞全部悬浮后, 加入对应体积的DMEM/1640完全培养基停止消化, 然后将其转移到离心管中进行离心。离心结束后弃掉上清, 加入新的DMEM/1640完全培养基重悬细胞, 最后转移到新的培养皿完成传代。
CCK-8测定    将处于对数生长期的细胞, 以每孔约3×103个细胞接种到96孔板中(每组3个复孔), 并置于37 ℃、5% CO2的培养箱中培养24 h, 然后用不同浓度的化合物处理(0、1.0、2.0、4.0、8.0、16.0、32.0、64.0 µmol·L-1), 培养24 h后避光向每孔加10 μL CCK-8溶液, 继续孵育60 min。用酶标仪在450 nm处测定吸光度值(OD), 计算出不同浓度下细胞的存活率, 绘制浓度-效应曲线, 并计算半抑制浓度(IC50)。细胞存活率(%) = (OD实验组-OD空白组)/(OD对照组-OD空白组) × 100%; 细胞抑制率(%) = (OD对照组-OD实验组)/(OD对照组-OD空白组) × 100%。
统计学分析    本文实验数据采用GraphPad Prism 9软件进行统计分析, 组间差异分析采用非配对Student's t检验比较, 数据采用平均值$ \stackrel{-}{x} $ ± s表示[15]
作者贡献: 任世杰和魏宏珂为本文共同第一作者, 负责化合物的分离、鉴定和文章撰写; 成欣欣、王晓敏和吕子玉对提取分离和结构鉴定提供了一定帮助; 王嘉琪和乔晓婷负责细胞相关实验、数据分析及作图; 曹朵负责实验的设计、稿件修改、对实验中遇到的问题给予指导等工作。
利益冲突: 所有作者均声明不存在利益冲突。
  • 陕西省自然科学基金(2024JC-YBQN-0920)
  • 陕西省科协技术学会青年人才托举计划项目(20220321)
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doi: 10.16438/j.0513-4870.2024-1190
  • 接收时间:2024-12-03
  • 首发时间:2025-10-29
  • 出版时间:2025-05-12
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  • 收稿日期:2024-12-03
  • 修回日期:2025-02-14
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陕西省自然科学基金(2024JC-YBQN-0920)
陕西省科协技术学会青年人才托举计划项目(20220321)
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    1.延安大学生命科学学院, 陕西省黄土高原资源植物研究与利用省市共建重点实验室, 陕西 延安 716000
    2.延安大学医学院, 陕西 延安 716000

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2种不同金属材料的力学参数

Family
属数
Number of
genus
种数
Number of
species
占总种数比例
Percentage of
total species (%)

Genus
种数
Number of
species
占总种数比例
Percentage of total
species (%)
鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
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