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Article(id=1195009886120559186, tenantId=1146029695717560320, journalId=1190317699101192196, issueId=1195009883369091469, articleNumber=1001-2494(2025)09-0956-10, orderNo=null, doi=10.11669/cpj.2025.09.008, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=research-article, receivedDate=1730304000000, receivedDateStr=2024-10-31, revisedDate=null, revisedDateStr=null, acceptedDate=null, acceptedDateStr=null, onlineDate=1762842159060, onlineDateStr=2025-11-11, pubDate=1746028800000, pubDateStr=2025-05-01, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1762842159060, onlineIssueDateStr=2025-11-11, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1762842159060, creator=13701087609, updateTime=1762842159060, updator=13701087609, issue=Issue{id=1195009883369091469, tenantId=1146029695717560320, journalId=1190317699101192196, year='2025', volume='60', issue='9', pageStart='893', pageEnd='1004', issueExtLink='null', onlineDate='null', pubDate='null', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=0, createTime=1762842158405, creator=13701087609, updateTime=1762846632399, updator=13701087609, preIssue=null, nextIssue=null, ext={EN=IssueExt(id=1195028649066893312, tenantId=1146029695717560320, journalId=1190317699101192196, issueId=1195009883369091469, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1195028649071087617, tenantId=1146029695717560320, journalId=1190317699101192196, issueId=1195009883369091469, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null}, startPage=956, endPage=965, ext={EN=ArticleExt(id=1195009886347051611, articleId=1195009886120559186, tenantId=1146029695717560320, journalId=1190317699101192196, language=EN, title=Quality Evaluation of Linderae Radix from Different Areas by Fingerprint Combined with Chemical Pattern Recognition, columnId=null, journalTitle=Chinese Pharmaceutical Journal, columnName=null, runingTitle=null, highlight=null, articleAbstract=

OBJECTIVE To establish HPLC fingerprint and multi-component content determination method for Linderae Radix, and explore the quality of Linderae Radix from different origins by combining chemical pattern recognition method, providing scientific basis for the quality control and further development of Linderae Radix. METHODS High-performance liquid chromatography(HPLC) was employed, using acetonitrile-0.1% triethylamine aqueous solution(adjusted to pH=3.0 with acetic acid) as the mobile phase for gradient elution, to establish the fingerprint of Linderae Radix. The quality of Linderae Radix from different areas was explored through chemical pattern recognition methods, such as similarity evaluation, cluster analysis(HCA), principal component analysis(PCA), and orthogonal partial least squares discriminant analysis(OPLS-DA). RESULTS The established fingerprint method meets the methodological requirements. A total of 17 common peaks were calibrated in the fingerprint spectra of 44 batches of Linderae Radix, and the similarity was greater than 0.8. Seven components were identified, including norisoboldine, boldine, reticuline, linderalactone, linderane, linderene and lindenanolide H. The HCA analysis results showed that the 44 batches of Linderae Radix were classified into five categories. PCA analysis indicated that the cumulative variance contribution rate of the first four principal components was 80.123%. OPLS-DA analysis showed that linderane, linderene, norisoboldine, and linderalactone were identified as differential chemical components of Linderae Radix. The content determination results indicate that “Tiantai Linderae Radix” has significantly higher levels of linderane (P<0.001), linderene (P<0.001), and linderalactone (P<0.01) compared with Linderae Radix from other regions. CONCLUSION The established fingerprint and content determination method is reliable and stable, providing a reference for the quality evaluation of Linderae Radix and the multi-index quality control of related preparations.

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目的 建立乌药(Linderae Radix)的高效液相色谱法(HPLC)指纹图谱及多成分含量测定方法,并结合化学模式识别法探究不同产地乌药的质量,为乌药的质量控制及进一步开发提供科学依据。方法 采用HPLC,以乙腈-体积分数0.1%的三乙胺水溶液(用乙酸调节pH=3.0)为流动相进行梯度洗脱,建立乌药指纹图谱,采用相似度评价、聚类分析(hierarchical cluster analysis,HCA)、主成分分析(principal component analysis,PCA)和正交偏最小二乘-判别分析(orthogonal partial least squares discriminant analysis,OPLS-DA)等化学模式识别法探讨不同产地乌药的质量。结果 建立的指纹图谱方法符合方法学要求,44批次乌药指纹图谱共标定17个共有峰,相似度均大于0.8,指认了去甲异波尔定,波尔定碱,瑞枯灵,乌药内酯,乌药醚内酯,乌药醇,lindenanolide H 7个成分;HCA将44批乌药分为5类;PCA得到前4个主成分的累计方差贡献率为80.123%;OPLS-DA筛选出乌药醚内酯,乌药醇,去甲异波尔定,乌药内酯为乌药差异性化学成分;含量测定结果表明,“天台乌药”在乌药醚内酯(P<0.001),乌药醇(P<0.001),乌药内酯(P<0.01)含量上显著高于其他产地乌药。结论 建立的指纹图谱和含量测定方法可靠、稳定,可为乌药药材质量评价及相关制剂的多指标质量控制提供参考。

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孙崇鲁,男,硕士,正高级实验师 研究方向:中药质量标准 Tel: (0574)88222743

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孙崇鲁,男,硕士,正高级实验师 研究方向:中药质量标准 Tel: (0574)88222743

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孙崇鲁,男,硕士,正高级实验师 研究方向:中药质量标准 Tel: (0574)88222743

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J Chin Pharm Sci, 2023, 45(2):123-130., articleTitle=Research progress on anti-hepatic injury effects of extracts of Lindera aggregata and its active components, refAbstract=null)], funds=[Fund(id=1195061446259254191, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, awardId=LGN21H280003, language=CN, fundingSource=浙江省公益基础研究项目资助(LGN21H280003), fundOrder=null, country=null), Fund(id=1195061446313780144, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, awardId=2019A610374, language=CN, fundingSource=宁波市自然基金项目资助(2019A610374), fundOrder=null, country=null)], companyList=[AuthorCompany(id=1195061441486136174, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, xref=null, ext=[AuthorCompanyExt(id=1195061441507107695, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, companyId=1195061441486136174, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=Zhejiang Pharmaceutical University, Ningbo 315100, China), AuthorCompanyExt(id=1195061441519690608, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, companyId=1195061441486136174, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=浙江药科职业大学, 浙江 宁波 315100)])], figs=[ArticleFig(id=1195061443985941397, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=EN, label=Fig.1, caption=HPLC superimposed fingerprint spectra of 44 batches of Linderae Radix, figureFileSmall=zaT/NIEg0GEICAJmPsQlGQ==, figureFileBig=4iAY4K51DkHJjGtZbhGjRg==, tableContent=null), ArticleFig(id=1195061444074021782, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=CN, label=图1, caption=44批乌药样品的高效液相色谱法(HPLC)叠加指纹图谱, figureFileSmall=zaT/NIEg0GEICAJmPsQlGQ==, figureFileBig=4iAY4K51DkHJjGtZbhGjRg==, tableContent=null), ArticleFig(id=1195061444220822423, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=EN, label=Fig.2, caption=HPLC diagram of mixed reference solution of Linderae Radix

5-norisoboldine; 6-boldine; 7-reticuline; 11-linderalactone; 12-linderane; 13-linderene; 16-lindenanolide H.

, figureFileSmall=g2gFxLn5a7ojCvJx2BGdcw==, figureFileBig=CNYstGFmmue+AzX4C/0wJQ==, tableContent=null), ArticleFig(id=1195061444296319896, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=CN, label=图2, caption=乌药样品混合对照品溶液的色谱图

5-去甲异波尔定;6-波尔定碱;7-瑞枯灵;11-乌药内酯;12-乌药醚内酯;13-乌药醇;16-lindenanolide H。

, figureFileSmall=g2gFxLn5a7ojCvJx2BGdcw==, figureFileBig=CNYstGFmmue+AzX4C/0wJQ==, tableContent=null), ArticleFig(id=1195061444363428761, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=EN, label=Fig.3, caption=Cluster analysis diagram of 44 batches of Linderae Radix, figureFileSmall=s4Nuu/tUmv6PnNbv4z3V0w==, figureFileBig=GaGuherqBktPK+qw/XDo9Q==, tableContent=null), ArticleFig(id=1195061444434731930, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=CN, label=图3, caption=44批乌药样品聚类分析图, figureFileSmall=s4Nuu/tUmv6PnNbv4z3V0w==, figureFileBig=GaGuherqBktPK+qw/XDo9Q==, tableContent=null), ArticleFig(id=1195061444497646491, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=EN, label=Fig.4, caption=OPLS-DA score plan of 44 batches of Linderae Radix samples, figureFileSmall=wjjKvTOiw1ZJ4Xogcej0tw==, figureFileBig=iVtmTTt2UJREWi6Hv8kkiw==, tableContent=null), ArticleFig(id=1195061444556366748, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=CN, label=图4, caption=44批乌药药材样品正交偏最小乘回归判别分析(OPLS-DA)得分平面图, figureFileSmall=wjjKvTOiw1ZJ4Xogcej0tw==, figureFileBig=iVtmTTt2UJREWi6Hv8kkiw==, tableContent=null), ArticleFig(id=1195061444657030045, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=EN, label=Fig.5, caption=Arrangement check diagram of OPLS-DA mode of Linderae Radix, figureFileSmall=e9Rbjb6r/2fXVEhCYMjQWQ==, figureFileBig=/WPSjgp4+d3VJVbx97k6bg==, tableContent=null), ArticleFig(id=1195061444724138910, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=CN, label=图5, caption=乌药样品OPLS-DA模型的排列检验图, figureFileSmall=e9Rbjb6r/2fXVEhCYMjQWQ==, figureFileBig=/WPSjgp4+d3VJVbx97k6bg==, tableContent=null), ArticleFig(id=1195061444791247775, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=EN, label=Fig.6, caption=VIP scores of 17 chromatographic peaks of Linderae Radix. n=6, figureFileSmall=oJcSzEP2UrUTlXZPRdhM8g==, figureFileBig=PtPB67dIgknlohxNFLD5Xw==, tableContent=null), ArticleFig(id=1195061444854162336, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=CN, label=图6, caption=乌药样品17个色谱峰的重要性投影值(VIP)得分图。n=6, figureFileSmall=oJcSzEP2UrUTlXZPRdhM8g==, figureFileBig=PtPB67dIgknlohxNFLD5Xw==, tableContent=null), ArticleFig(id=1195061444992574369, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=EN, label=Tab.1, caption=

Information of Linderae Radix samples from different places at different times

, figureFileSmall=null, figureFileBig=null, tableContent=
No. Source(in Chinese) Collection time No. Source(in Chinese) Collection time
S1 Tiantai county, Zhejiang Province(浙江省天台县) 2021-07-22 S23 Tiantai county, Zhejiang Province(浙江省天台县) 2023-08-19
S2 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S24 Tiantai county, Zhejiang province(浙江省天台县) 2023-08-19
S3 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S25 Tiantai county, Zhejiang province(浙江省天台县) 2023-08-19
S4 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S26 Suichang county, Zhejiang province(浙江省遂昌县) 2022-08-07
S5 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S27 Longyou county, Zhejiang province(浙江省龙游县) 2022-08-07
S6 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S28 Hengshan county, Hunan province(湖南省衡山县) 2021-07-23
S7 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S29 Hengshan county, Hunan province(湖南省衡山县) 2021-07-23
S8 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S30 Yueyang city, Hunan province(湖南省岳阳市) 2021-07-24
S9 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S31 Yueyang city, Hunan province(湖南省岳阳市) 2021-07-24
S10 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S32 Yongzhou city, Hunan province(湖南省永州市) 2021-07-24
S11 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S33 Yongzhou city, Hunan province(湖南省永州市) 2021-07-24
S12 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S34 Xiajiang county, Jiangxi province(江西省峡江县) 2022-08-07
S13 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S35 Xiajiang county, Jiangxi province(江西省峡江县) 2022-08-07
S14 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S36 Shangrao city, Jiangxi province(江西省上饶市) 2022-08-07
S15 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S37 Yichun city, Jiangxi province(江西省宜春市) 2022-08-08
S16 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S38 Yichun city, Jiangxi province(江西省宜春市) 2022-08-08
S17 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S39 Zhangshu city, Jiangxi province(江西省樟树市) 2022-08-08
S18 Tiantai county, Zhejiang province(浙江省天台县) 2023-08-19 S40 Zhangshu city, Jiangxi province(江西省樟树市) 2022-08-08
S19 Tiantai county, Zhejiang province(浙江省天台县) 2023-08-19 S41 Huangshan city, Anhui province(安徽省黄山市) 2022-09-13
S20 Tiantai county, Zhejiang province(浙江省天台县) 2023-08-19 S42 Huangshan city, Anhui province(安徽省黄山市) 2022-09-13
S21 Tiantai county, Zhejiang province(浙江省天台县) 2023-08-19 S43 Shaoguan city, Guangdong province(广东省韶关市) 2020-06-18
S22 Tiantai county, Zhejiang province(浙江省天台县) 2023-08-19 S44 Shaoguan city, Guangdong province(广东省韶关市) 2020-06-18
), ArticleFig(id=1195061445072266146, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=CN, label=表1, caption=

44批次乌药样品产地信息

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No. Source(in Chinese) Collection time No. Source(in Chinese) Collection time
S1 Tiantai county, Zhejiang Province(浙江省天台县) 2021-07-22 S23 Tiantai county, Zhejiang Province(浙江省天台县) 2023-08-19
S2 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S24 Tiantai county, Zhejiang province(浙江省天台县) 2023-08-19
S3 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S25 Tiantai county, Zhejiang province(浙江省天台县) 2023-08-19
S4 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S26 Suichang county, Zhejiang province(浙江省遂昌县) 2022-08-07
S5 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S27 Longyou county, Zhejiang province(浙江省龙游县) 2022-08-07
S6 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S28 Hengshan county, Hunan province(湖南省衡山县) 2021-07-23
S7 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S29 Hengshan county, Hunan province(湖南省衡山县) 2021-07-23
S8 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S30 Yueyang city, Hunan province(湖南省岳阳市) 2021-07-24
S9 Tiantai county, Zhejiang province(浙江省天台县) 2021-07-22 S31 Yueyang city, Hunan province(湖南省岳阳市) 2021-07-24
S10 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S32 Yongzhou city, Hunan province(湖南省永州市) 2021-07-24
S11 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S33 Yongzhou city, Hunan province(湖南省永州市) 2021-07-24
S12 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S34 Xiajiang county, Jiangxi province(江西省峡江县) 2022-08-07
S13 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S35 Xiajiang county, Jiangxi province(江西省峡江县) 2022-08-07
S14 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S36 Shangrao city, Jiangxi province(江西省上饶市) 2022-08-07
S15 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S37 Yichun city, Jiangxi province(江西省宜春市) 2022-08-08
S16 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S38 Yichun city, Jiangxi province(江西省宜春市) 2022-08-08
S17 Tiantai county, Zhejiang province(浙江省天台县) 2022-08-06 S39 Zhangshu city, Jiangxi province(江西省樟树市) 2022-08-08
S18 Tiantai county, Zhejiang province(浙江省天台县) 2023-08-19 S40 Zhangshu city, Jiangxi province(江西省樟树市) 2022-08-08
S19 Tiantai county, Zhejiang province(浙江省天台县) 2023-08-19 S41 Huangshan city, Anhui province(安徽省黄山市) 2022-09-13
S20 Tiantai county, Zhejiang province(浙江省天台县) 2023-08-19 S42 Huangshan city, Anhui province(安徽省黄山市) 2022-09-13
S21 Tiantai county, Zhejiang province(浙江省天台县) 2023-08-19 S43 Shaoguan city, Guangdong province(广东省韶关市) 2020-06-18
S22 Tiantai county, Zhejiang province(浙江省天台县) 2023-08-19 S44 Shaoguan city, Guangdong province(广东省韶关市) 2020-06-18
), ArticleFig(id=1195061445177123747, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=EN, label=Tab.2, caption=

Characteristic values and contribution rate of principal components in principal component analysis of Linderae Radix

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Principal
component		 Initial eigenvalue
Characteristic value Variance contribution rate/% Cumulative variance contribution rate/%
1 6.670 39.235 39.235
2 3.791 22.299 61.534
3 2.069 12.171 73.705
4 1.091 6.418 80.123
), ArticleFig(id=1195061445286175652, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=CN, label=表2, caption=

乌药样品主成分分析主成分特征值及贡献率

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Principal
component		 Initial eigenvalue
Characteristic value Variance contribution rate/% Cumulative variance contribution rate/%
1 6.670 39.235 39.235
2 3.791 22.299 61.534
3 2.069 12.171 73.705
4 1.091 6.418 80.123
), ArticleFig(id=1195061445391033253, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=EN, label=Tab.3, caption=

Rotated component matrix for principal component analysis of Linderae Radix

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Peak
No.		 Principal component Peak
No.		 Principal component
L1 L2 L3 L4 L1 L2 L3 L4
1 0.936 -0.126 0.166 0.049 10 0.391 -0.072 0.767 0.038
2 0.961 -0.117 0.079 -0.076 11 -0.097 0.912 0.01 0.031
3 0.945 -0.048 0.128 -0.119 12 0.028 0.726 0.393 0.176
4 0.963 -0.183 0.033 -0.082 13 -0.131 0.895 0.109 0.115
5 0.683 -0.018 0.029 -0.406 14 -0.156 0.871 -0.019 -0.137
6 0.328 0.525 -0.12 -0.121 15 -0.193 0.061 -0.085 0.919
7 0.971 -0.06 0.164 -0.022 14 -0.193 0.773 -0.391 -0.059
8 0.062 -0.146 0.794 0.041 17 -0.384 0.716 -0.256 0.235
9 0.01 0.153 0.832 -0.226
), ArticleFig(id=1195061445470725030, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=CN, label=表3, caption=

乌药样品主成分分析旋转后的成分矩阵

, figureFileSmall=null, figureFileBig=null, tableContent=
Peak
No.		 Principal component Peak
No.		 Principal component
L1 L2 L3 L4 L1 L2 L3 L4
1 0.936 -0.126 0.166 0.049 10 0.391 -0.072 0.767 0.038
2 0.961 -0.117 0.079 -0.076 11 -0.097 0.912 0.01 0.031
3 0.945 -0.048 0.128 -0.119 12 0.028 0.726 0.393 0.176
4 0.963 -0.183 0.033 -0.082 13 -0.131 0.895 0.109 0.115
5 0.683 -0.018 0.029 -0.406 14 -0.156 0.871 -0.019 -0.137
6 0.328 0.525 -0.12 -0.121 15 -0.193 0.061 -0.085 0.919
7 0.971 -0.06 0.164 -0.022 14 -0.193 0.773 -0.391 -0.059
8 0.062 -0.146 0.794 0.041 17 -0.384 0.716 -0.256 0.235
9 0.01 0.153 0.832 -0.226
), ArticleFig(id=1195061445533639591, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=EN, label=Tab.4, caption=

Calibration curves, correlation coefficients, linear ranges of seven components of Linderae Radix

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Component Regression equation r Range of linearity/μg·mL-1
Norisoboldine y=33 361x+13 330 0.999 2 7.22 -288.68
Boldine y=36 755x-15 402 0.999 9 2.02 -80.88
Reticuline y=20 106x+17 737 0.999 8 5.54 -221.40
Linderalactone y=6 882.3x+14 046 0.999 8 4.77 -190.06
Linderane y=11 206x+67 350 0.999 9 3.23 -129.12
Linderene y=7 851.6x+7 653.5 0.999 8 6.83 -273.36
Lindenanolide H y=5 072.3x-3 633.2 0.999 6 2.06 -82.32
), ArticleFig(id=1195061445592359848, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=CN, label=表4, caption=

乌药中7种成分的回归方程、相关系数、线性范围

, figureFileSmall=null, figureFileBig=null, tableContent=
Component Regression equation r Range of linearity/μg·mL-1
Norisoboldine y=33 361x+13 330 0.999 2 7.22 -288.68
Boldine y=36 755x-15 402 0.999 9 2.02 -80.88
Reticuline y=20 106x+17 737 0.999 8 5.54 -221.40
Linderalactone y=6 882.3x+14 046 0.999 8 4.77 -190.06
Linderane y=11 206x+67 350 0.999 9 3.23 -129.12
Linderene y=7 851.6x+7 653.5 0.999 8 6.83 -273.36
Lindenanolide H y=5 072.3x-3 633.2 0.999 6 2.06 -82.32
), ArticleFig(id=1195061445676245929, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=EN, label=Tab.5, caption=

Results of recovery tests for 7 components in Linderae Radix. n=6

, figureFileSmall=null, figureFileBig=null, tableContent=
Component m(Sample)/g m(Original)/mg m(Added)/mg m(Measured)/mg Recovery/% Average recovery/% RSD/%
Norisoboldine 0.201 1 0.551 0 0.600 0 1.157 2 101.03 100.32 1.57
0.203 2 0.556 8 0.600 0 1.162 1 100.89
0.202 2 0.554 0 0.600 0 1.147 9 98.98
0.201 4 0.551 8 0.600 0 1.145 4 98.93
0.203 8 0.558 4 0.600 0 1.175 8 102.90
0.201 7 0.552 7 0.600 0 1.147 9 99.21
Boldine 0.201 3 0.149 0 0.160 0 0.311 8 101.78 101.82 1.66
0.203 3 0.150 4 0.160 0 0.308 8 98.97
0.205 4 0.152 0 0.160 0 0.316 3 102.66
0.201 9 0.149 4 0.160 0 0.312 4 101.86
0.201 5 0.149 1 0.160 0 0.311 5 101.51
0.204 3 0.151 2 0.160 0 0.317 8 104.12
Reticuline 0.205 5 0.096 6 0.100 0 0.193 4 96.84 100.79 2.65
0.201 4 0.094 7 0.100 0 0.196 8 102.09
0.200 1 0.094 0 0.100 0 0.197 2 103.18
0.201 1 0.094 5 0.100 0 0.196 8 102.24
0.200 9 0.094 4 0.100 0 0.192 4 97.96
0.201 7 0.094 8 0.100 0 0.197 2 102.43
Linderalactone 0.199 8 0.917 1 1.000 0 1.910 9 99.38 98.36 1.96
0.202 2 0.928 1 1.000 0 1.919 4 99.13
0.205 3 0.942 3 1.000 0 1.900 4 95.81
0.198 9 0.913 0 1.000 0 1.916 4 100.34
0.199 3 0.914 8 1.000 0 1.909 2 99.44
0.202 5 0.929 5 1.000 0 1.889 9 96.05
Linderane 0.201 2 0.484 9 0.400 0 0.892 1 101.79 100.66 1.26
0.201 4 0.485 4 0.400 0 0.882 1 99.19
0.198 7 0.478 9 0.400 0 0.874 5 98.91
0.199 1 0.479 8 0.400 0 0.885 3 101.37
0.199 2 0.480 1 0.400 0 0.886 4 101.59
0.200 7 0.483 7 0.400 0 0.888 1 101.10
Linderene 0.203 3 1.462 7 1.450 0 2.895 5 98.82 98.34 0.44
0.201 6 1.450 4 1.450 0 2.880 7 98.64
0.201 7 1.451 1 1.450 0 2.879 3 98.49
0.200 8 1.444 7 1.450 0 2.867 8 98.15
0.199 2 1.433 2 1.450 0 2.859 1 98.34
0.200 7 1.444 0 1.450 0 2.859 2 97.60
Lindenanolide H 0.202 2 0.491 3 0.500 0 1.012 1 104.15 102.36 1.72
0.198 8 0.483 1 0.500 0 0.997 5 102.88
0.203 2 0.493 8 0.500 0 0.998 7 100.98
0.201 8 0.490 4 0.500 0 0.993 7 100.67
0.199 4 0.484 5 0.500 0 1.007 6 104.61
0.202 7 0.492 6 0.500 0 0.997 0 100.89
), ArticleFig(id=1195061445760132010, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=CN, label=表5, caption=

乌药中7个成分的加样回收率试验结果。n=6

, figureFileSmall=null, figureFileBig=null, tableContent=
Component m(Sample)/g m(Original)/mg m(Added)/mg m(Measured)/mg Recovery/% Average recovery/% RSD/%
Norisoboldine 0.201 1 0.551 0 0.600 0 1.157 2 101.03 100.32 1.57
0.203 2 0.556 8 0.600 0 1.162 1 100.89
0.202 2 0.554 0 0.600 0 1.147 9 98.98
0.201 4 0.551 8 0.600 0 1.145 4 98.93
0.203 8 0.558 4 0.600 0 1.175 8 102.90
0.201 7 0.552 7 0.600 0 1.147 9 99.21
Boldine 0.201 3 0.149 0 0.160 0 0.311 8 101.78 101.82 1.66
0.203 3 0.150 4 0.160 0 0.308 8 98.97
0.205 4 0.152 0 0.160 0 0.316 3 102.66
0.201 9 0.149 4 0.160 0 0.312 4 101.86
0.201 5 0.149 1 0.160 0 0.311 5 101.51
0.204 3 0.151 2 0.160 0 0.317 8 104.12
Reticuline 0.205 5 0.096 6 0.100 0 0.193 4 96.84 100.79 2.65
0.201 4 0.094 7 0.100 0 0.196 8 102.09
0.200 1 0.094 0 0.100 0 0.197 2 103.18
0.201 1 0.094 5 0.100 0 0.196 8 102.24
0.200 9 0.094 4 0.100 0 0.192 4 97.96
0.201 7 0.094 8 0.100 0 0.197 2 102.43
Linderalactone 0.199 8 0.917 1 1.000 0 1.910 9 99.38 98.36 1.96
0.202 2 0.928 1 1.000 0 1.919 4 99.13
0.205 3 0.942 3 1.000 0 1.900 4 95.81
0.198 9 0.913 0 1.000 0 1.916 4 100.34
0.199 3 0.914 8 1.000 0 1.909 2 99.44
0.202 5 0.929 5 1.000 0 1.889 9 96.05
Linderane 0.201 2 0.484 9 0.400 0 0.892 1 101.79 100.66 1.26
0.201 4 0.485 4 0.400 0 0.882 1 99.19
0.198 7 0.478 9 0.400 0 0.874 5 98.91
0.199 1 0.479 8 0.400 0 0.885 3 101.37
0.199 2 0.480 1 0.400 0 0.886 4 101.59
0.200 7 0.483 7 0.400 0 0.888 1 101.10
Linderene 0.203 3 1.462 7 1.450 0 2.895 5 98.82 98.34 0.44
0.201 6 1.450 4 1.450 0 2.880 7 98.64
0.201 7 1.451 1 1.450 0 2.879 3 98.49
0.200 8 1.444 7 1.450 0 2.867 8 98.15
0.199 2 1.433 2 1.450 0 2.859 1 98.34
0.200 7 1.444 0 1.450 0 2.859 2 97.60
Lindenanolide H 0.202 2 0.491 3 0.500 0 1.012 1 104.15 102.36 1.72
0.198 8 0.483 1 0.500 0 0.997 5 102.88
0.203 2 0.493 8 0.500 0 0.998 7 100.98
0.201 8 0.490 4 0.500 0 0.993 7 100.67
0.199 4 0.484 5 0.500 0 1.007 6 104.61
0.202 7 0.492 6 0.500 0 0.997 0 100.89
), ArticleFig(id=1195061445869183915, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=EN, label=Tab.6, caption=

Determination results of 7 indicator components in 44 batches of Linderae Radix. mg·g-1

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Sample No. Norisoboldine Boldine Reticuline Linderalactone Linderane Linderene Lindenanolide H Total
S1 4.46 0.91 2.29 3.73 3.19 8.60 1.81 24.99
S2 3.25 0.63 0.94 4.94 5.67 12.95 2.45 30.82
S3 5.86 1.07 3.12 2.51 2.00 6.37 1.90 22.83
S4 4.07 0.81 2.44 4.50 4.04 11.58 2.18 29.61
S5 4.73 0.86 2.87 1.90 2.88 4.10 0.35 17.70
S6 2.74 0.74 0.47 4.59 2.41 7.20 2.43 20.57
S7 5.62 1.05 3.66 2.53 3.19 6.35 0.36 22.76
S8 2.36 0.58 1.11 2.19 2.53 5.71 1.50 15.98
S9 2.05 0.26 1.32 2.37 2.31 6.22 0.99 15.52
S10 4.45 0.80 2.56 2.61 1.79 5.77 0.65 18.63
S11 4.58 0.55 4.40 1.36 1.61 3.58 0.25 16.34
S12 5.18 1.06 7.82 2.53 2.39 6.55 0.35 25.87
S13 5.82 0.18 4.75 2.36 2.33 5.34 0.25 21.03
S14 2.45 0.45 1.51 2.14 1.77 4.85 0.83 14.01
S15 4.37 0.54 0.83 4.33 4.02 9.78 1.72 25.59
S16 5.47 2.94 4.20 5.44 3.83 11.57 6.50 39.94
S17 2.36 0.82 1.88 2.88 1.92 6.38 0.29 16.54
S18 4.00 0.49 4.35 4.04 3.96 11.77 0.53 29.13
S19 4.50 0.53 1.78 1.26 2.76 4.05 0.96 15.85
S20 3.34 1.33 2.84 3.24 1.96 3.89 0.26 16.85
S21 10.10 1.10 10.06 3.55 3.71 5.29 0.55 34.36
S22 8.35 2.29 6.99 3.86 4.36 9.31 0.47 35.64
S23 5.19 0.96 6.40 3.78 3.57 9.55 0.47 29.92
S24 2.42 1.08 1.00 5.82 3.58 10.93 1.83 26.66
S25 7.52 1.75 5.04 3.99 4.18 10.59 0.58 33.66
S26 3.64 1.53 3.25 3.38 1.58 8.15 2.31 23.83
S27 2.43 0.82 2.42 1.48 0.46 2.97 0.13 10.70
S28 2.69 0.52 0.85 2.08 0.84 3.40 0.32 10.70
S29 4.50 0.76 2.33 1.95 0.38 3.23 0.61 13.77
S30 12.11 0.48 5.73 1.45 0.38 2.97 0.88 24.00
S31 5.89 0.94 6.50 1.30 1.04 1.27 0.20 17.14
S32 5.49 0.66 5.49 1.55 1.33 1.55 0.13 16.19
S33 5.14 1.22 6.86 1.25 1.25 1.79 0.22 17.73
S34 4.91 0.51 2.17 2.27 3.01 6.11 2.02 21.01
S35 5.86 0.47 2.46 5.21 0.76 6.04 2.99 23.80
S36 3.29 0.99 1.38 1.06 0.92 2.73 0.56 10.93
S37 8.36 0.51 4.66 1.03 2.23 1.16 0.19 18.14
S38 9.34 0.72 3.11 3.06 1.05 4.93 1.65 23.86
S39 6.16 0.30 1.18 2.59 2.64 4.77 1.11 18.76
S40 8.01 0.14 4.30 1.07 2.27 1.87 0.17 17.84
S41 6.26 0.45 2.71 0.66 1.36 2.21 0.43 14.08
S42 5.69 0.57 2.15 3.28 1.32 5.71 1.87 20.59
S43 8.97 0.40 3.27 3.09 1.32 6.39 2.08 25.52
S44 9.38 0.45 6.38 1.43 1.63 3.65 0.18 23.11
), ArticleFig(id=1195061445978235820, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=CN, label=表6, caption=

不同产地乌药7个指标性成分含量测定结果。mg·g-1

, figureFileSmall=null, figureFileBig=null, tableContent=
Sample No. Norisoboldine Boldine Reticuline Linderalactone Linderane Linderene Lindenanolide H Total
S1 4.46 0.91 2.29 3.73 3.19 8.60 1.81 24.99
S2 3.25 0.63 0.94 4.94 5.67 12.95 2.45 30.82
S3 5.86 1.07 3.12 2.51 2.00 6.37 1.90 22.83
S4 4.07 0.81 2.44 4.50 4.04 11.58 2.18 29.61
S5 4.73 0.86 2.87 1.90 2.88 4.10 0.35 17.70
S6 2.74 0.74 0.47 4.59 2.41 7.20 2.43 20.57
S7 5.62 1.05 3.66 2.53 3.19 6.35 0.36 22.76
S8 2.36 0.58 1.11 2.19 2.53 5.71 1.50 15.98
S9 2.05 0.26 1.32 2.37 2.31 6.22 0.99 15.52
S10 4.45 0.80 2.56 2.61 1.79 5.77 0.65 18.63
S11 4.58 0.55 4.40 1.36 1.61 3.58 0.25 16.34
S12 5.18 1.06 7.82 2.53 2.39 6.55 0.35 25.87
S13 5.82 0.18 4.75 2.36 2.33 5.34 0.25 21.03
S14 2.45 0.45 1.51 2.14 1.77 4.85 0.83 14.01
S15 4.37 0.54 0.83 4.33 4.02 9.78 1.72 25.59
S16 5.47 2.94 4.20 5.44 3.83 11.57 6.50 39.94
S17 2.36 0.82 1.88 2.88 1.92 6.38 0.29 16.54
S18 4.00 0.49 4.35 4.04 3.96 11.77 0.53 29.13
S19 4.50 0.53 1.78 1.26 2.76 4.05 0.96 15.85
S20 3.34 1.33 2.84 3.24 1.96 3.89 0.26 16.85
S21 10.10 1.10 10.06 3.55 3.71 5.29 0.55 34.36
S22 8.35 2.29 6.99 3.86 4.36 9.31 0.47 35.64
S23 5.19 0.96 6.40 3.78 3.57 9.55 0.47 29.92
S24 2.42 1.08 1.00 5.82 3.58 10.93 1.83 26.66
S25 7.52 1.75 5.04 3.99 4.18 10.59 0.58 33.66
S26 3.64 1.53 3.25 3.38 1.58 8.15 2.31 23.83
S27 2.43 0.82 2.42 1.48 0.46 2.97 0.13 10.70
S28 2.69 0.52 0.85 2.08 0.84 3.40 0.32 10.70
S29 4.50 0.76 2.33 1.95 0.38 3.23 0.61 13.77
S30 12.11 0.48 5.73 1.45 0.38 2.97 0.88 24.00
S31 5.89 0.94 6.50 1.30 1.04 1.27 0.20 17.14
S32 5.49 0.66 5.49 1.55 1.33 1.55 0.13 16.19
S33 5.14 1.22 6.86 1.25 1.25 1.79 0.22 17.73
S34 4.91 0.51 2.17 2.27 3.01 6.11 2.02 21.01
S35 5.86 0.47 2.46 5.21 0.76 6.04 2.99 23.80
S36 3.29 0.99 1.38 1.06 0.92 2.73 0.56 10.93
S37 8.36 0.51 4.66 1.03 2.23 1.16 0.19 18.14
S38 9.34 0.72 3.11 3.06 1.05 4.93 1.65 23.86
S39 6.16 0.30 1.18 2.59 2.64 4.77 1.11 18.76
S40 8.01 0.14 4.30 1.07 2.27 1.87 0.17 17.84
S41 6.26 0.45 2.71 0.66 1.36 2.21 0.43 14.08
S42 5.69 0.57 2.15 3.28 1.32 5.71 1.87 20.59
S43 8.97 0.40 3.27 3.09 1.32 6.39 2.08 25.52
S44 9.38 0.45 6.38 1.43 1.63 3.65 0.18 23.11
), ArticleFig(id=1195061446049538989, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=EN, label=Tab.7, caption=

Mann-Whitney U test results for 7 components in Tiantai vs other regions' Linderae Radix

, figureFileSmall=null, figureFileBig=null, tableContent=
Component Content(Average±SD)/mg·g-1 U-value P-value
Group A(S1-S25, n=25) Group B(S26-S44, n=19)
Norisoboldine 4.61±1.96 6.22±2.57 135.00 0.015
Boldine 0.95±0.61 0.63±0.33 148.50 0.035
Reticuline 3.39±2.43 3.54±1.89 214.00 0.578
Linderalactone 3.30±1.23 2.06±1.13 103.00 0.001
Linderane 3.04±1.03 1.36±0.74 37.00 <0.001
Linderene 7.53±2.87 3.73±2.02 63.50 <0.001
Lindenanolide H 0.98±0.75 0.95±0.91 189.00 0.250
), ArticleFig(id=1195061446120842158, tenantId=1146029695717560320, journalId=1190317699101192196, articleId=1195009886120559186, language=CN, label=表7, caption=

天台乌药与其他产地乌药7种成分含量的Mann-Whitney U检验结果

, figureFileSmall=null, figureFileBig=null, tableContent=
Component Content(Average±SD)/mg·g-1 U-value P-value
Group A(S1-S25, n=25) Group B(S26-S44, n=19)
Norisoboldine 4.61±1.96 6.22±2.57 135.00 0.015
Boldine 0.95±0.61 0.63±0.33 148.50 0.035
Reticuline 3.39±2.43 3.54±1.89 214.00 0.578
Linderalactone 3.30±1.23 2.06±1.13 103.00 0.001
Linderane 3.04±1.03 1.36±0.74 37.00 <0.001
Linderene 7.53±2.87 3.73±2.02 63.50 <0.001
Lindenanolide H 0.98±0.75 0.95±0.91 189.00 0.250
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基于指纹图谱结合化学模式识别法评价不同产地乌药的质量
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孙崇鲁 , 方琳 , 孙平飞 , 岳军治 , 刘京骅
中国药学杂志 | 论著 2025,60(9): 956-965
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中国药学杂志 | 论著 2025, 60(9): 956-965
基于指纹图谱结合化学模式识别法评价不同产地乌药的质量
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孙崇鲁, 方琳, 孙平飞, 岳军治, 刘京骅
作者信息
  • 浙江药科职业大学, 浙江 宁波 315100

    • 孙崇鲁,男,硕士,正高级实验师 研究方向:中药质量标准 Tel: (0574)88222743

Quality Evaluation of Linderae Radix from Different Areas by Fingerprint Combined with Chemical Pattern Recognition
Chonglu SUN, Lin FANG, Pingfei SUN, Junzhi YUE, Jinghua LIU
Affiliations
  • Zhejiang Pharmaceutical University, Ningbo 315100, China
出版时间: 2025-05-01 doi: 10.11669/cpj.2025.09.008
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目的 建立乌药(Linderae Radix)的高效液相色谱法(HPLC)指纹图谱及多成分含量测定方法,并结合化学模式识别法探究不同产地乌药的质量,为乌药的质量控制及进一步开发提供科学依据。方法 采用HPLC,以乙腈-体积分数0.1%的三乙胺水溶液(用乙酸调节pH=3.0)为流动相进行梯度洗脱,建立乌药指纹图谱,采用相似度评价、聚类分析(hierarchical cluster analysis,HCA)、主成分分析(principal component analysis,PCA)和正交偏最小二乘-判别分析(orthogonal partial least squares discriminant analysis,OPLS-DA)等化学模式识别法探讨不同产地乌药的质量。结果 建立的指纹图谱方法符合方法学要求,44批次乌药指纹图谱共标定17个共有峰,相似度均大于0.8,指认了去甲异波尔定,波尔定碱,瑞枯灵,乌药内酯,乌药醚内酯,乌药醇,lindenanolide H 7个成分;HCA将44批乌药分为5类;PCA得到前4个主成分的累计方差贡献率为80.123%;OPLS-DA筛选出乌药醚内酯,乌药醇,去甲异波尔定,乌药内酯为乌药差异性化学成分;含量测定结果表明,“天台乌药”在乌药醚内酯(P<0.001),乌药醇(P<0.001),乌药内酯(P<0.01)含量上显著高于其他产地乌药。结论 建立的指纹图谱和含量测定方法可靠、稳定,可为乌药药材质量评价及相关制剂的多指标质量控制提供参考。

乌药  /  指纹图谱  /  化学模式识别  /  含量测定  /  质量控制

OBJECTIVE To establish HPLC fingerprint and multi-component content determination method for Linderae Radix, and explore the quality of Linderae Radix from different origins by combining chemical pattern recognition method, providing scientific basis for the quality control and further development of Linderae Radix. METHODS High-performance liquid chromatography(HPLC) was employed, using acetonitrile-0.1% triethylamine aqueous solution(adjusted to pH=3.0 with acetic acid) as the mobile phase for gradient elution, to establish the fingerprint of Linderae Radix. The quality of Linderae Radix from different areas was explored through chemical pattern recognition methods, such as similarity evaluation, cluster analysis(HCA), principal component analysis(PCA), and orthogonal partial least squares discriminant analysis(OPLS-DA). RESULTS The established fingerprint method meets the methodological requirements. A total of 17 common peaks were calibrated in the fingerprint spectra of 44 batches of Linderae Radix, and the similarity was greater than 0.8. Seven components were identified, including norisoboldine, boldine, reticuline, linderalactone, linderane, linderene and lindenanolide H. The HCA analysis results showed that the 44 batches of Linderae Radix were classified into five categories. PCA analysis indicated that the cumulative variance contribution rate of the first four principal components was 80.123%. OPLS-DA analysis showed that linderane, linderene, norisoboldine, and linderalactone were identified as differential chemical components of Linderae Radix. The content determination results indicate that “Tiantai Linderae Radix” has significantly higher levels of linderane (P<0.001), linderene (P<0.001), and linderalactone (P<0.01) compared with Linderae Radix from other regions. CONCLUSION The established fingerprint and content determination method is reliable and stable, providing a reference for the quality evaluation of Linderae Radix and the multi-index quality control of related preparations.

Linderae Radix  /  fingerprint  /  chemical pattern recognition  /  content determination  /  quality control
孙崇鲁, 方琳, 孙平飞, 岳军治, 刘京骅. 基于指纹图谱结合化学模式识别法评价不同产地乌药的质量. 中国药学杂志, 2025 , 60 (9) : 956 -965 . DOI: 10.11669/cpj.2025.09.008
Chonglu SUN, Lin FANG, Pingfei SUN, Junzhi YUE, Jinghua LIU. Quality Evaluation of Linderae Radix from Different Areas by Fingerprint Combined with Chemical Pattern Recognition[J]. Chinese Pharmaceutical Journal, 2025 , 60 (9) : 956 -965 . DOI: 10.11669/cpj.2025.09.008
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乌药[Lindera aggregata (Sims) Kosterm]为樟科山胡椒属植物,2018年遴选为新“浙八味”之一。乌药主要产于浙江、江西、福建、湖南、安徽、广东等省区[1-2]。乌药以块根入药,具有行气止痛、温肾散寒等功效,乌药在民间和临床上药用广泛,作为多种中药复方和中成药的重要药味,比如木香理气片、妇科宁坤丸、开郁顺气丸和解表追风丸等[3-5]。现代药理学研究表明乌药具有抗炎镇痛、抗肿瘤、抗氧化等广泛的药理活性[6-8]。浙江是乌药的主产区,占全国产量的70%以上。传统认为浙江天台为乌药著名的道地产区,称之为“天台乌药”或“台乌药”。近年来对乌药化学成分研究发现其化学成分主要包括萜类及内酯类、生物碱类、挥发油类等[9-11]。目前关于乌药质量评价方法仅以一种或一类化学成分为指标[12-14],难以实现对乌药药材道地性的全面评价。本研究以不同产地乌药为研究对象,通过高效液相色谱法(HPLC)建立乌药指纹图谱,结合化学模式识别法对共有峰面积数据进行分析,以实现对乌药的综合评价[15-16],并建立乌药多成分含量测定方法,以期为乌药药材的质量评价、质量控制及道地性提供方法和参考依据。
1 材料
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1.1 仪器
高效液相色谱仪(LC-20AT型,日本岛津公司);超声波清洗仪(SB25-12D 型,宁波新艺超声设备有限公司);十万分之一电子分析天平(AUW120D型,日本岛津公司);电子天平(ME104E型,梅特勒-托利多仪器有限公司);精密恒温水浴锅(J-HH-6A型,上海冠森生物科技公司)。
1.2 试剂与药材
对照品去甲异波尔定(批号:J12HB184948),波尔定碱(批号:X23N9Y73113),瑞枯灵(批号:J06IB205338),乌药内酯(批号:R01N9F74005),乌药醚内酯(批号:M16J12S137719),乌药醇(批号:Z02N9S74009)(上海源叶生物科技有限公司);Lindenanolide H为乌药叶提取分离纯化所得,经电喷雾-质谱(ESI-MS)、核磁共振氢谱(1H-NMR)和核磁共振碳谱(13C-NMR)确认,HPLC测定化合物纯度,归一化法计算纯度>95%。乙腈(色谱纯,TEDIA公司),甲醇、冰乙酸、二氯甲烷、乙醇、三乙胺等(分析纯,国药集团化学试剂有限公司),液相色谱流动相用水为娃哈哈纯净水。乌药药材采自浙江、湖南、江西、安徽、广东等,共44批次。经宁波中药研究院彭昕教授鉴定为乌药的干燥根茎,其产地信息见表1。
2 方法与结果
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2.1 色谱条件
采用Inertsil ODS-SP C18(4.6 mm×150 mm,5.0 μm)色谱柱,流动相:乙腈(A)-体积分数0.1%的三乙胺水溶液(用乙酸调节pH=3)(B),二元高压梯度洗脱0.1→15 min,10%→20%A;15→25 min,20→60%A;25→40 min,60%→70%A;40→55 min,70%→100%A;55~60 min 10%A;检测波长235 nm;柱温:40 ℃;总流速:0.900 mL·min-1;进样量:10 μL。
2.2 溶液的制备
2.2.1 对照品溶液制备
精密称定各对照品去甲异波尔定,波尔定碱,瑞枯灵,乌药内酯,乌药醚内酯,乌药醇,lindenanolide H适量,分别于5 mL量瓶中,加入适量的甲醇溶解并定容,制得各对照品浓度分别为2.062、2.022、2.214、1.906、2.152、2.278、2.058 mg·mL-1的单一对照品溶液。精密吸取上述单一对照品溶液各适量于同一20 mL量瓶中,用甲醇定容至刻度后摇匀,得到去甲异波尔定,波尔定碱,瑞枯灵,乌药内酯,乌药醚内酯,乌药醇,lindenanolide H质量浓度分别为360.850、101.100、276.750、238.250、161.400、341.700、102.900 μg·mL-1的混合对照品溶液,于4 ℃条件下保存,备用。
2.2.2 供试品溶液制备
精密称取乌药粉末约0.2 g,置50 mL具塞锥形瓶中,精密加入甲醇10 mL,密塞,称定重量,超声处理(功率500 W,频率40 kHz)60 min,放冷,再称定重量,用甲醇补足减失的重量,摇匀,滤过,取续滤液过0.22 μm的微孔滤膜,即得供试品溶液。
2.3 乌药指纹图谱的建立
2.3.1 精密度试验
取乌药粉末(S6),按“2.2.2”项下描述的方法制备溶液,按色谱分析方法“2.1”项下描述的条件,连续进样6次,以去甲异波尔定为标准参照峰(R),计算得到17个共有峰相对保留时间的相对标准偏差(RSD)在0.01%~0.07%,相对峰面积的RSD在0.43%~2.96%,表明仪器精密度良好。
2.3.2 重复性试验
取同一乌药粉末(S6),按供试品溶液制备方法“2.2.2”项下描述的方法平行制备6份,分别按色谱分析方法“2.1”项下描述的条件进样测定,以去甲异波尔定为参照峰(R),计算得到17个共有峰相对保留时间的RSD在0.66%~3.51%,相对峰面积的RSD在0.66%~3.51%,表明方法重复性好。
2.3.3 稳定性试验
取乌药粉末(S6),按“2.2.2”项下描述的方法制备溶液,在制备后的不同时间点(即 0、2、4、6、12和24 h)按色谱分析方法“2.1”项下描述的条件进行分析,以去甲异波尔定为参照峰,计算得到17个共有峰相对保留时间的RSD在0.01%~0.09%,相对峰面积的RSD在0.47%~4.23%,表明供试品溶液在24 h内稳定性良好。
2.3.4 指纹图谱相似度评价及共有峰指认
将44批乌药按“2.2.2”项下描述的方法制备成供试品溶液,然后分别按色谱分析方法“2.1”项下条件分析,将所得色谱数据文件导入到“中药色谱指纹图谱相似度评价系统软件”,将S1设置为对照图谱,时间窗宽度设置为 0.18 min,采用中位数法生成44批乌药药材的HPLC指纹图谱和对照图谱(R),结果见图1。最终确定不同产地乌药有17个共有峰,通过与对照品图谱对比,共指认了7个主要特征峰,其中峰5为去甲异波尔定、峰6为波尔定碱、峰7为瑞枯灵、峰11为乌药内酯、峰12为乌药醚内酯、峰13为乌药醇、峰16为lindenanolide H,见图2。44批乌药色谱图中特征共有峰的相对保留时间的RSD<0.5%,但是共有峰面积的RSD差异较大,说明不同产地各成分含量有显著不同,这可能与乌药属于广分布品种,不同的生长环境、生长年限等因素影响植物的次生代谢。44批样品与对照图谱比较,相似度评价结果均大于0.8,表明不同产地乌药质量相对稳定,可为乌药药材的品质评价提供科学依据。
2.4 化学模式识别分析
2.4.1 聚类分析(HCA)
采用IBM SPSS Statistics 26统计软件,以44批乌药17个共有峰面积为变量,采用组间联接法进行HCA,具体见图3。 当分类距离为10时,显示44批的乌药一共可聚为5类,其中S30、S37、S38、S40、S43、S44聚为一类,S21聚为一类,S16、S22、S25聚为一类,S2、S6、S8、S9、S14、S17、S24、S27、S28、S36聚为一类,其他剩余样品共同聚为一类。说明不同产地乌药的化学成分存在差异,药材质量与产地有一定的关联,但是不同产地的药材在聚类时也有一定的交叉。这可能是由于不同产地的乌药在生长年限、药材基源和生长环境等方面存在差异,进而影响了其次生代谢产物的累积[17]。
2.4.2 主成分分析(PCA)
以17个共有峰的峰面积数值为变量,采用IBM SPSS Statistics 26软件进行PCA,结果显示抽样适合性检验(Kaiser-Meyer-Olkin,KMO)值为0.736(>0.5),表明变量相关性较好。以特征值大于1为提取标准,得到前4个主成分因子(L1, L2, L3和L4)的特征值分别为6.670、3.791、2.069、1.091。前4个主成分累积方差贡献率为80.123%(表2),这说明了主成分因子1~4能基本上反映乌药的整体信息。将初始因子载荷矩阵进行正交旋转,得到17个共有峰成分在4个主成分中的旋转成分矩阵,见表3。旋转成分矩阵显示了不同共有峰对4个主成分不同方差贡献率,其绝对值越大说明该成分在决定药材区分中的作用越大[18]。因此,第1个主成分信息主要来自峰7(瑞枯灵)、峰4 、峰2、峰3、峰1、峰5 (去甲异波尔定);第2个主成分信息主要来自峰11(乌药内酯)、峰13(乌药醇)、峰14、峰16(lindenanolide H)、峰12(乌药醚内酯)、峰17、峰6(波尔定碱);第3个主成分信息主要来自峰8 、峰9 、峰10;第4个主成分信息主要来自峰15。
2.4.3 正交偏最小二乘-判别分析(OPLS-DA)
为了更好地体现乌药质量差异,将44批不同产地乌药指纹图谱中17个共有峰峰面积数值导入SIMAC14.1软件,进行OPLS-DA建模分析,其中散点得分图见图4。R2和Q2分别代表模型可解释的变量和可预测度,可对模型的优劣进行判别,理论上R2、Q2数值越接近1,说明模型越好,越低说明模型的拟合准确性越差,通常情况下,R2、Q2高于0.5较好[19]。该模型的R2x=0.683,R2Y=0.762,Q2=0.651,均大于0.5,说明模型的拟合效果好,模型稳定可靠。结果显示44批乌药样品被分为2类,浙江地区的为一类,其他产地为一类。将建立的模型通过200次置换检验,结果见图5。从图中可以看出,R2和Q2的回归线斜率均大于0,这表明随着模型复杂度的增加,模型在新样本上的解释能力和预测性能都在逐步提高。R2和Q2的回归线截距在OPLS-DA模型中分别代表了模型在没有自变量时的基线拟合能力和预测能力。这两个截距的大小对于评估模型的复杂度、拟合度和预测性能具有重要意义[20]。此模型的R2和Q2回归线截距为0.157和-0.55,说明建立的OPLS-DA 模型未出现过拟合,预测能力较好且有效可用,能够对不同产地的乌药进行判别分析。在OPLS-DA模型中,变量重要性投影值(variable importance in the projection,VIP)是评估不同自变量对因变量的贡献程度,是筛选差异性化合物的重要指标[21]。以VIP>1为条件筛选出差异标志物,由VIP得分图(图6)可知,对乌药样品的影响较大4个差异化合物依次为12号峰(乌药醚内酯)>13号峰(乌药醇)>5号峰(去甲异波尔定)>11号峰(乌药内酯),其对应的 VIP值依次为 1.511、1.432、1.295、1.182。
2.5 7个指标性成分的含量测定
2.5.1 线性关系的考察
分别移取“2.2.1”项下混合对照品溶液0.1、0.2、0.5、1.0、2.5、4.0 mL置5 mL的量瓶中,用甲醇定容后摇匀,得到梯度浓度的混合对照品溶液,按“2.1”项下描述的色谱条件进样分析,记录峰面积。然后将浓度作为自变量(X),峰面积作为因变量(Y),并应用线性回归分析,得相关结果见表4。
2.5.2 精密度试验
取混合对照品储备液,按“2.1”项下的色谱分析条件连续进样6次,结果去甲异波尔定,波尔定碱,瑞枯灵,乌药内酯,乌药醚内酯,乌药醇,lindenanolide H保留时间的RSD均小于0.02%,峰面积的RSD分别为0.83%、1.13%、0.79%、1.17%、1.44%、1.06%、1.11%,表明仪器精密度良好。
2.5.3 重复性试验
精密称取0.2 g乌药粉末(S6) 6份,按供试品溶液制备方法“2.2.2”项下方法制备6份溶液,按色谱分析方法“2.1”项下的条件进样分析,结果去甲异波尔定,波尔定碱,瑞枯灵,乌药内酯,乌药醚内酯,乌药醇,lindenanolide H保留时间的RSD均小于0.05%,峰面积的RSD分别为3.08%、2.70%、3.30%、3.35%、3.51%、1.68%、2.96%,表明方法重复性良好。
2.5.4 稳定性试验
精密称取乌药粉末(S6)0.2g,按供试品溶液制备方法“2.2.2”项下方法制备供试品溶液,在制备后的不同时间点(即 0、2、4、6、12和24 h)进行色谱分析。 结果显示,去甲异波尔定,波尔定碱,瑞枯灵,乌药内酯,乌药醚内酯,乌药醇,lindenanolide H保留时间的RSD均为0.01%,峰面积的 RSD分别为 1.58%、0.85%、0.65%、0.78%、0.56%、0.47%、1.76%,表明样品24 h内稳定性良好。
2.5.5 回收率试验
精密称定已测质量浓度乌药粉末(S6) 6份,每份0.2g,加入适量对照品。按供试品溶液制备方法“2.2.2”项下方法制备供试品溶液,按色谱分析方法“2.1”项下条件进行分析,根据线性回归方程得出化合物含量,并计算回收率,结果见表5。结果显示,去甲异波尔定的平均回收率为100.32%,RSD为1.57%;波尔定碱的平均回收率为101.82%,RSD为1.66%,瑞枯灵的平均回收率为100.79%,RSD为2.65%,乌药内酯的平均回收率为98.36%,RSD为1.96%,乌药醚内酯的平均回收率为100.66%,RSD为1.26%,乌药醇的平均回收率为98.34%,RSD为0.44%,lindenanolide H的平均回收率为102.36%,RSD为1.72%,回收率介于98.34%~102.36%,RSD均小于3%。
2.5.6 样品含量测定
待测样品溶液按上述色谱方法进行分析,根据线性回归方程计算出化合物含量,测定结果见表6。
为了探究不同产地乌药在成分含量上的差异性,本研究采用IBM SPSS Statistics 26统计软件进行分析。由于本研究中样本数据不满足正态分布或方差齐性等参数检验的前提条件,因此选用非参数Mann-Whitney U检验方法,分析结果见表7。由表7可知,“天台乌药”与其他产地乌药中乌药醚内酯和乌药醇含量呈现极显著差异(P <0.001),乌药内酯含量呈现高度显著差异(P <0.01),去甲异波尔定和波尔定碱含量呈现显著差异(P<0.05),瑞枯灵和lindenanolide H含量差异不显著(P>0.05)。Mann-Whitney U检验表明,“天台乌药”在多个活性成分含量上显著高于其他产地乌药,尤其是在乌药醚内酯、乌药醇和乌药内酯等成分上表现出极显著或高度显著差异。这些结果为“天台乌药”的药用价值提供了科学依据,并为进一步研究其成分差异的成因奠定了基础。
3 讨论
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本实验对色谱条件进行了系统考察,包括不同溶剂(二氯甲烷、乙腈、体积分数70%乙醇、体积分数95%乙醇、体积分数95%甲醇、体积分数70%甲醇)对乌药指纹图谱提取效果的影响,以指纹图谱色谱峰数量、峰型和分离度为指标,最终确定甲醇作为提取溶剂;其次考察了4种流动相组成(乙腈-水、甲醇-水、乙腈-体积分数0.05%磷酸水溶液、乙腈-体积分数0.1%三乙胺醋酸水溶液)对供试品溶液的分离效果,结果发现以乙腈-0.1%三乙胺水溶液(用乙酸调节pH=3.0)为流动相时,样品的色谱峰峰型较好且分离度最佳;考察了不同波长(220、235、275、320 nm)下指纹图谱色谱峰数量、响应值和分离度,结果发现235 nm下各色谱峰的响应较高而且分离度较好,因此选用235 nm为色谱检测波长。另外也考察了洗脱程序、柱温、流速等对样品的分离效果的影响,最终确定了色谱条件。
本实验利用HPLC建立了44批乌药药材指纹图谱,相似度评价结果表明,44批乌药药材相似度均大于0.8,但共有峰面积RSD差异较大,说明不同乌药药材批次整体相似度较高,但是成分含量差异较大。不同产地乌药样品共有17个共有峰,利用对照品指认了其中7个峰,分别为去甲异波尔定,波尔定碱,瑞枯灵,乌药内酯,乌药醚内酯,乌药醇,lindenanolide H。HCA结果显示,当分类距离为10时,将44批样品聚成5类,不同省份的乌药在聚类上有明显交叉。这可能与生长年限、药材基源、日照等密切相关,同时气候环境、经纬度、海拔、土壤等影响也会导致成分含量上的变化[22]。PCA结果显示,前4个主成分累积方差贡献率为80.123%。OPLS-DA结果显示,44批乌药样品被分为2类,浙江地区的为一类,其他产地为一类。通过模型VIP值发现乌药醚内酯,乌药醇,去甲异波尔定,乌药内酯是乌药的差异性特征成分,可以作为区分和鉴别不同产区乌药药材质量标志物。
含量测定结果显示,44批次样品中7种成分含量差异较大。其中,去甲异波尔定含量为2.05~12.11 mg·g-1,波尔定碱0.14~2.94 mg·g-1,瑞枯灵为0.47~10.06 mg·g-1,乌药内酯为0.66~5.82 mg·g-1,乌药醚内酯为0.38~5.67 mg·g-1,乌药醇为1.16~12.95 mg·g-1,lindenanolide H为 0.13~6.5 mg·g-1,7种成分的总含量为10.7~39.94 mg·g-1。平均含量上从高到低依次为:乌药醇>去甲异波尔定>瑞枯灵>乌药内酯>乌药醚内酯>lindenanolide H>波尔定碱。进一步对比“天台乌药”和其他产地乌药中上述成分的含量,发现“天台乌药”中乌药醚内酯、乌药内酯和乌药醇的含量显著高于其他产地,这可能与其独特的生长环境和遗传背景有关。现代药理学研究表明,乌药醚内酯,乌药醇,乌药内酯等具有多种生物活性和药理作用,其中乌药醚内酯具有镇痛、抗炎、抗疲劳、保肝、抗菌抗病毒等药理活性[23],乌药内酯具有抗菌抗病毒、抗炎、抗氧化、抗肿瘤、心血管保护等药理活性[24],乌药醇具有保肝、抗菌抗病毒、抗肿瘤、心血管保护等药理活性[25]。因此,乌药醚内酯、乌药醇和乌药内酯的含量变化对乌药的整体质量和药效有着重要影响。这也进一步表明,乌药醚内酯、乌药内酯和乌药醇可能作为“天台乌药”的标志性成分,为其道地性提供了重要依据。
本研究建立的HPLC指纹图谱、化学模式识别方法以及多成分含量测定方法稳定、可靠,为乌药药材的质量控制和相关资源开发利用提供参考价值。“天台乌药”中乌药醚内酯、乌药醇和乌药内酯的高含量,为其道地性提供了重要依据。后续研究将进一步结合代谢组学和基因组学技术,深入分析乌药中的未知成分,以期为乌药药材的品质评价及其道地性研究提供更为全面的科学依据。
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  • 浙江省公益基础研究项目资助(LGN21H280003)
  • 宁波市自然基金项目资助(2019A610374)
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2025年第60卷第9期
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doi: 10.11669/cpj.2025.09.008
  • 接收时间：2024-10-31
  • 首发时间：2025-11-11
  • 出版时间：2025-05-01
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  • 收稿日期：2024-10-31
基金
浙江省公益基础研究项目资助(LGN21H280003)
宁波市自然基金项目资助(2019A610374)
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    浙江药科职业大学, 浙江 宁波 315100
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2种不同金属材料的力学参数
科
Family		 属数
Number of
genus		 种数
Number of
species		 占总种数比例
Percentage of
total species (%)		 属
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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