Article(id=1239173808922349915, tenantId=1146029695717560320, journalId=1205117023404326918, issueId=1239173808419033435, articleNumber=null, orderNo=null, doi=10.16155/j.0254-1793.2024.02.16, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=null, receivedDate=1687104000000, receivedDateStr=2023-06-19, revisedDate=null, revisedDateStr=null, acceptedDate=null, acceptedDateStr=null, onlineDate=1773371658783, onlineDateStr=2026-03-13, pubDate=1709136000000, pubDateStr=2024-02-29, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1773371658783, onlineIssueDateStr=2026-03-13, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1773371658783, creator=13701087609, updateTime=1773371658783, updator=13701087609, issue=Issue{id=1239173808419033435, tenantId=1146029695717560320, journalId=1205117023404326918, year='2024', volume='44', issue='2', pageStart='185', pageEnd='372', issueExtLink='null', onlineDate='null', pubDate='null', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=null, createTime=1773371658663, creator=13701087609, updateTime=1773371757717, updator=13701087609, preIssue=null, nextIssue=null, ext={EN=IssueExt(id=1239174223944536397, tenantId=1146029695717560320, journalId=1205117023404326918, issueId=1239173808419033435, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1239174223944536398, tenantId=1146029695717560320, journalId=1205117023404326918, issueId=1239173808419033435, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null}, startPage=324, endPage=332, ext={EN=ArticleExt(id=1239173809174008158, articleId=1239173808922349915, tenantId=1146029695717560320, journalId=1205117023404326918, language=EN, title=Polygonati Rhizoma polysaccharide differential analysis based on enzymolysis and chemometrics*, columnId=1239173809106899293, journalTitle=Chinese Journal of Pharmaceutical Analysis, columnName=Quality Contro, runingTitle=null, highlight=null, articleAbstract=
Objective:

To determine the polysaccharide hydrolysates from Polygonati Rhizoma under glycoside enzymatic hydrolysis’ HPLC fingerprint. Examine the variations among the polysaccharides produced by the various varieties of Polygonati Rhizoma and provide references for the assessment of the polysaccharide quality.

Methods:

After the polysaccharide from Polygonati Rhizoma was hydrolyzed by fructose enzymes, its fingerprint was established by HPLC-ELSD. The fingerprint was then analyzed using similarity analysis (SA), hierarchical cluster analysis (HCA), and principal component analysis (PCA) to determine the differences between the polysaccharides from various Polygonati Rhizoma varieties.

Results:

Polysaccharides from various strains of Polygonati Rhizoma had different HPLC-ELSD fingerprints, and a total of 17 distinct oligosaccharide fragments were discovered, all of which contained fructose, glucose, and sucrose. Analysis revealed that there are significant intra species differences, minor differences, and high levels of similarity between the three types of Polygonati Rhizoma. The real Polygonati Rhizoma differs significantly from the imitation in several important ways.

Conclusion:

Polygonati Rhizoma can be successfully classified according to varieties, using the fingerprint of polysaccharide hydrolysates of the various varieties of Polygonati Rhizoma and its adulterants. The developed HPLC method can be used for the differential analysis of polysaccharides in Polygonati Rhizoma and is straightforward, precise, and repeatable.

, correspAuthors=Ya-zhong ZHANG, authorNote=null, correspAuthorsNote=null, copyrightStatement=null, 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=Zhuo DIAO, Chong HU, Qing-shan YANG, Ya-zhong ZHANG), CN=ArticleExt(id=1239173811015307670, articleId=1239173808922349915, tenantId=1146029695717560320, journalId=1205117023404326918, language=CN, title=基于酶解法和化学计量学的黄精多糖差异性分析*, columnId=1239148842025799861, journalTitle=药物分析杂志, columnName=质量分析, runingTitle=null, highlight=null, articleAbstract=
目的:

建立糖苷酶水解下不同品种黄精多糖酶解产物的HPLC指纹图谱,探讨不同品种黄精多糖的差异,为黄精药材多糖质量评价提供参考。

方法:

黄精多糖经果糖苷酶水解后,采用HPLC建立黄精的指纹图谱;并结合相似度(SA)分析、聚类分析(HCA)和主成分分析(PCA),对指纹图谱进行分析,研究不同品种黄精多糖的差异。

结果:

不同品种黄精的多糖HPLC指纹图谱存在差异,共得到17个特异性寡糖片段,均含有果糖、葡萄糖和蔗糖。分析表明,3种基原黄精的种内指纹色谱相似度较高,差异较小,种间差异较大,而黄精混伪品与基原黄精差异明显。

结论:

不同品种黄精及其混伪品多糖酶解产物的指纹图谱结合化学计量学分析可以成功将黄精按照品种进行分类。所建立的HPLC方法简便、准确,重复性好,可用于黄精药材的多糖的差异研究。

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**Tel:13956985695;E-mail:
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Tel:18356875022;E-mail:

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J Chromatogr A20111218(51): 922, articleTitle=Characterizing plant cell wall derived oligosaccharides using hydrophilic interaction chromatography with mass spectrometry detection, refAbstract=null)], funds=[Fund(id=1239173819341001402, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, awardId=null, language=CN, fundingSource=*安徽省药品监督管理局监管科学研究重点项目-常用中药材DNA分子鉴定研究, fundOrder=null, country=null)], companyList=[AuthorCompany(id=1239173811304714656, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, xref=1., ext=[AuthorCompanyExt(id=1239173811313103265, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, companyId=1239173811304714656, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=1.Anhui Medical College, Hefei 230031, China), AuthorCompanyExt(id=1239173811321491874, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, companyId=1239173811304714656, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=1.安徽医学高等专科学校,合肥 230031)]), AuthorCompany(id=1239173811405377959, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, xref=2., ext=[AuthorCompanyExt(id=1239173811413766568, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, companyId=1239173811405377959, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=2.Anhui Provincial Institute for Food and Drug Control, Key Laboratory of Quality Research and Evaluation of Traditional Chinese Medicines of the State Food and Drug Administration, Hefei 230051, China), AuthorCompanyExt(id=1239173811422155177, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, companyId=1239173811405377959, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=2.安徽省食品药品检验研究院 国家药监局中药质量研究与评价重点实验室,合肥 230051)]), AuthorCompany(id=1239173812894355885, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, xref=3., ext=[AuthorCompanyExt(id=1239173812902744495, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, companyId=1239173812894355885, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=3.Anhui University of Chinese Medicine, Hefei 230012, China), AuthorCompanyExt(id=1239173812911133103, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, companyId=1239173812894355885, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=3.安徽中医药大学,合肥 230012)])], figs=[ArticleFig(id=1239173815549350455, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, language=EN, label=Fig.1, caption=Overlay map of control map of Polygonati Rhizoma polysaccharide from different varieties, figureFileSmall=7QkFAfi/5oeb24GlbA0oIg==, figureFileBig=/o0aMgIcLX+YkoQsjDi0SQ==, tableContent=null), ArticleFig(id=1239173815620653627, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, language=CN, label=图1, caption=不同品种黄精多糖对照图谱叠加图

1.果糖(fructose) 2.葡萄糖(glucose) 4.蔗糖(sucrose)

, figureFileSmall=7QkFAfi/5oeb24GlbA0oIg==, figureFileBig=/o0aMgIcLX+YkoQsjDi0SQ==, tableContent=null), ArticleFig(id=1239173815872311875, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, language=EN, label=Fig.2, caption=Polysaccharide fingerprint of 10 batches of Polygonatum cyrtonema Hua, figureFileSmall=SETMAyzgr5yex3+G+Mz23g==, figureFileBig=A9MN0HZyL8L3z4f5NpyvIw==, tableContent=null), ArticleFig(id=1239173817336123981, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, language=CN, label=图2, caption=10批多花黄精多糖指纹图谱

R.多花黄精对照图谱(the control chromatogram of Polygonatum cyrtonema Hua) S1-S10.多花黄精(Polygonatum cyrtonema Hua)

, figureFileSmall=SETMAyzgr5yex3+G+Mz23g==, figureFileBig=A9MN0HZyL8L3z4f5NpyvIw==, tableContent=null), ArticleFig(id=1239173817478730320, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, language=EN, label=Fig.3, caption=Fingerprint of Polygonatum cyrtonema Hua polysaccharides as reference, figureFileSmall=cvplSSMH4w/KQqQ+MGaiwA==, figureFileBig=UJrt3sJlAYv1S6t5UlcZHg==, tableContent=null), ArticleFig(id=1239173817575199320, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, language=CN, label=图3, caption=以多花黄精多糖为参考不同品种黄精的指纹图谱

R1.多花黄精对照图谱(the control chromagram of Polygonatum cyrtonema Hua) D1~D5.滇黄精(Polygonatum kingianum Coll.et Hemsl.) J1~J5.黄精(Polygonatum sibiricum Red) H1~H5.湖北黄精(Polygonatum zanlanscianense Pamp) C1~C5.四川黄精(Sichuan Polygonati Rhizoma) Z1~Z2.进口黄精(imported Polygonati Rhizoma) X1~X2.小叶黄精(Polygonati Rhizoma microphylla)

, figureFileSmall=cvplSSMH4w/KQqQ+MGaiwA==, figureFileBig=UJrt3sJlAYv1S6t5UlcZHg==, tableContent=null), ArticleFig(id=1239173817671668316, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, language=EN, label=Fig.4, caption=HCA dendrogram relationship of 20 batches of Polygonati Rhizoma polysaccharide, figureFileSmall=v6oXpusoAsWeQkM4gOH5OA==, figureFileBig=3QgwsWpB8snjsK8GYeBpaw==, tableContent=null), ArticleFig(id=1239173817776525921, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, language=CN, label=图4, caption=20批黄精多糖HCA树状关系

1-10.多花黄精(Polygonatum cyrtonema Hua) 11-15.滇黄精(Polygonatum kingianum Coll. et) 16-20.黄精(Polygonatum sibiricum Red)

, figureFileSmall=v6oXpusoAsWeQkM4gOH5OA==, figureFileBig=3QgwsWpB8snjsK8GYeBpaw==, tableContent=null), ArticleFig(id=1239173817868800616, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, language=EN, label=Fig.5, caption=Scree plot of 20 batches of Polygonati Rhizoma polysaccharide, figureFileSmall=WN29/QZ85x2qcJsQlH9X/w==, figureFileBig=tYnhOFe0D7OfWa0dLuEthw==, tableContent=null), ArticleFig(id=1239173817952686700, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, language=CN, label=图5, caption=20批黄精多糖碎石图, figureFileSmall=WN29/QZ85x2qcJsQlH9X/w==, figureFileBig=tYnhOFe0D7OfWa0dLuEthw==, tableContent=null), ArticleFig(id=1239173818032378481, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, language=EN, label=Fig.6, caption=PCA 3D projection of 20 batches of Polygonati Rhizoma polysaccharide, figureFileSmall=u0PGWm6aBUuE6YyDYKG6ww==, figureFileBig=1MBWYyscIti/8gP8ORGODg==, tableContent=null), ArticleFig(id=1239173818120458872, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, language=CN, label=图6, caption=20批黄精多糖PCA三维投影

1.多花黄精(Polygonatum cyrtonema Hua) 2.滇黄精(Polygonatum kingianum Coll.et Hemsl.) 3.黄精(Polygonatum sibiricum Red)

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1.多花黄精(Polygonatum cyrtonema Hua) 2.滇黄精(Polygonatum kingianum Coll.et Hemsl.) 3.鸡头黄精(Polygonatum sibiricum Red)

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M1-M17.峰1~17(peak 1-17)

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Detailed list of test materials

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编号
(number)
产地
(habitat)
编号
(number)
产地
(habitat)
S1江西瑞昌(Ruichang,Jiangxi)D3贵州毕节(Bijie,Guizhou)
S2安徽六安(Lu’an,Anhui)D4云南昆明(Kunming,Yunnan)
S3安徽池州(Chizhou,Anhui)D5四川达州(Dazhou,Sichuan)
S4安徽池州(Chizhou,Anhui)C1四川成都(Chengdu,Sichuan)
S5安徽池州(Chizhou,Anhui)C2湖北武汉(Wuhan,Hubei)
S6浙江杭州(Hangzhou,Zhejiang)C3河南洛阳(Luoyang,Henan)
S7浙江杭州(Hangzhou,Zhejiang)C4河南洛阳(Luoyang,Henan)
S8浙江杭州(Hangzhou,Zhejiang)C5河南洛阳(Luoyang,Henan)
S9浙江杭州(Hangzhou,Zhejiang)H1湖南岳阳(Yueyang,Hunan)
S10浙江衢州(Quzhou,Zhejiang)H2湖南岳阳(Yueyang,Hunan)
J1贵州毕节(Bijie,Guizhou)H3安徽池州(Chizhou,Anhui)
J2安徽池州(Chizhou,Anhui)H4安徽池州(Chizhou,Anhui)
J3山东泰安(Tai’an,Shandong)H5安徽池州(Chizhou,Anhui)
J4辽宁抚顺(Fushun,Liaoning)Z1国外(abroad)
J5河南洛阳(Luoyang,Henan)Z2国外(abroad)
D1四川成都(Chengdu,Sichuan)X1四川成都(Chengdu,Sichuan)
D2湖北武汉(Wuhan,Hubei)X2四川成都(Chengdu,Sichuan)
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试验材料详表

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编号
(number)
产地
(habitat)
编号
(number)
产地
(habitat)
S1江西瑞昌(Ruichang,Jiangxi)D3贵州毕节(Bijie,Guizhou)
S2安徽六安(Lu’an,Anhui)D4云南昆明(Kunming,Yunnan)
S3安徽池州(Chizhou,Anhui)D5四川达州(Dazhou,Sichuan)
S4安徽池州(Chizhou,Anhui)C1四川成都(Chengdu,Sichuan)
S5安徽池州(Chizhou,Anhui)C2湖北武汉(Wuhan,Hubei)
S6浙江杭州(Hangzhou,Zhejiang)C3河南洛阳(Luoyang,Henan)
S7浙江杭州(Hangzhou,Zhejiang)C4河南洛阳(Luoyang,Henan)
S8浙江杭州(Hangzhou,Zhejiang)C5河南洛阳(Luoyang,Henan)
S9浙江杭州(Hangzhou,Zhejiang)H1湖南岳阳(Yueyang,Hunan)
S10浙江衢州(Quzhou,Zhejiang)H2湖南岳阳(Yueyang,Hunan)
J1贵州毕节(Bijie,Guizhou)H3安徽池州(Chizhou,Anhui)
J2安徽池州(Chizhou,Anhui)H4安徽池州(Chizhou,Anhui)
J3山东泰安(Tai’an,Shandong)H5安徽池州(Chizhou,Anhui)
J4辽宁抚顺(Fushun,Liaoning)Z1国外(abroad)
J5河南洛阳(Luoyang,Henan)Z2国外(abroad)
D1四川成都(Chengdu,Sichuan)X1四川成都(Chengdu,Sichuan)
D2湖北武汉(Wuhan,Hubei)X2四川成都(Chengdu,Sichuan)
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Similarity of different varieties of Polygonati Rhizoma based on the comparison map of Polygonatum cyrtonema Hua polysaccharide

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样品
(sample)
相似度
(similarity)
样品
(sample)
相似度
(similarity)
S10.937D30.827
S20.998D40.779
S30.997D50.723
S40.997C10.762
S50.998C20.858
S60.997C30.824
S70.998C40.901
S80.998C50.694
S90.997H10.896
S100.997H20.877
J10.755H30.932
J20.838H40.913
J30.770H50.445
J40.751Z10.765
J50.343Z20.818
D10.933X10.445
D20.773X20.474
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基于多花黄精多糖对照图谱的不同品种黄精相似度

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样品
(sample)
相似度
(similarity)
样品
(sample)
相似度
(similarity)
S10.937D30.827
S20.998D40.779
S30.997D50.723
S40.997C10.762
S50.998C20.858
S60.997C30.824
S70.998C40.901
S80.998C50.694
S90.997H10.896
S100.997H20.877
J10.755H30.932
J20.838H40.913
J30.770H50.445
J40.751Z10.765
J50.343Z20.818
D10.933X10.445
D20.773X20.474
), ArticleFig(id=1239173819034817190, tenantId=1146029695717560320, journalId=1205117023404326918, articleId=1239173808922349915, language=EN, label=Tab.3, caption=

Characteristic value and variance contribution rate

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成分
(principal component)
初始特征值(initial eigenvalue)提取载荷平方和(extraction sums of squared loadings)
总计
(total)
方差百分比
(variance percentage)
累积
(accumulate)
总计
(total)
方差百分比
(variance percentage)
累积
(accumulate)
18.52250.12950.1298.52250.12950.129
24.51926.58076.7094.51926.58076.709
31.5539.13585.8441.5539.13585.844
41.1296.63892.4821.1296.63892.482
50.5433.19395.675
60.3171.86797.542
70.1440.84998.391
80.1020.59998.990
90.0780.45799.447
100.0420.24899.695
110.0200.11699.811
120.0170.10099.911
130.0070.03899.949
140.0040.02599.974
150.0030.01899.992
160.0010.00799.999
179.277×10-50.001100.000
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特征值和方差贡献率

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成分
(principal component)
初始特征值(initial eigenvalue)提取载荷平方和(extraction sums of squared loadings)
总计
(total)
方差百分比
(variance percentage)
累积
(accumulate)
总计
(total)
方差百分比
(variance percentage)
累积
(accumulate)
18.52250.12950.1298.52250.12950.129
24.51926.58076.7094.51926.58076.709
31.5539.13585.8441.5539.13585.844
41.1296.63892.4821.1296.63892.482
50.5433.19395.675
60.3171.86797.542
70.1440.84998.391
80.1020.59998.990
90.0780.45799.447
100.0420.24899.695
110.0200.11699.811
120.0170.10099.911
130.0070.03899.949
140.0040.02599.974
150.0030.01899.992
160.0010.00799.999
179.277×10-50.001100.000
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基于酶解法和化学计量学的黄精多糖差异性分析*
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刁卓 1 , 胡冲 2 , 杨青山 3 , 张亚中 2, 3, **
药物分析杂志 | 质量分析 2024,44(2): 324-332
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药物分析杂志 | 质量分析 2024, 44(2): 324-332
基于酶解法和化学计量学的黄精多糖差异性分析*
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刁卓1 , 胡冲2, 杨青山3, 张亚中2, 3, **
作者信息
  • 1.安徽医学高等专科学校,合肥 230031
  • 2.安徽省食品药品检验研究院 国家药监局中药质量研究与评价重点实验室,合肥 230051
  • 3.安徽中医药大学,合肥 230012
  • Tel:18356875022;E-mail:

通讯作者:

**Tel:13956985695;E-mail:
Polygonati Rhizoma polysaccharide differential analysis based on enzymolysis and chemometrics*
Zhuo DIAO1 , Chong HU2, Qing-shan YANG3, Ya-zhong ZHANG2, 3, **
Affiliations
  • 1.Anhui Medical College, Hefei 230031, China
  • 2.Anhui Provincial Institute for Food and Drug Control, Key Laboratory of Quality Research and Evaluation of Traditional Chinese Medicines of the State Food and Drug Administration, Hefei 230051, China
  • 3.Anhui University of Chinese Medicine, Hefei 230012, China
出版时间: 2024-02-29 doi: 10.16155/j.0254-1793.2024.02.16
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目的:

建立糖苷酶水解下不同品种黄精多糖酶解产物的HPLC指纹图谱,探讨不同品种黄精多糖的差异,为黄精药材多糖质量评价提供参考。

方法:

黄精多糖经果糖苷酶水解后,采用HPLC建立黄精的指纹图谱;并结合相似度(SA)分析、聚类分析(HCA)和主成分分析(PCA),对指纹图谱进行分析,研究不同品种黄精多糖的差异。

结果:

不同品种黄精的多糖HPLC指纹图谱存在差异,共得到17个特异性寡糖片段,均含有果糖、葡萄糖和蔗糖。分析表明,3种基原黄精的种内指纹色谱相似度较高,差异较小,种间差异较大,而黄精混伪品与基原黄精差异明显。

结论:

不同品种黄精及其混伪品多糖酶解产物的指纹图谱结合化学计量学分析可以成功将黄精按照品种进行分类。所建立的HPLC方法简便、准确,重复性好,可用于黄精药材的多糖的差异研究。

黄精  /  多糖  /  酶解法  /  指纹图谱  /  差异  /  化学计量学  /  高效液相色谱法
Objective:

To determine the polysaccharide hydrolysates from Polygonati Rhizoma under glycoside enzymatic hydrolysis’ HPLC fingerprint. Examine the variations among the polysaccharides produced by the various varieties of Polygonati Rhizoma and provide references for the assessment of the polysaccharide quality.

Methods:

After the polysaccharide from Polygonati Rhizoma was hydrolyzed by fructose enzymes, its fingerprint was established by HPLC-ELSD. The fingerprint was then analyzed using similarity analysis (SA), hierarchical cluster analysis (HCA), and principal component analysis (PCA) to determine the differences between the polysaccharides from various Polygonati Rhizoma varieties.

Results:

Polysaccharides from various strains of Polygonati Rhizoma had different HPLC-ELSD fingerprints, and a total of 17 distinct oligosaccharide fragments were discovered, all of which contained fructose, glucose, and sucrose. Analysis revealed that there are significant intra species differences, minor differences, and high levels of similarity between the three types of Polygonati Rhizoma. The real Polygonati Rhizoma differs significantly from the imitation in several important ways.

Conclusion:

Polygonati Rhizoma can be successfully classified according to varieties, using the fingerprint of polysaccharide hydrolysates of the various varieties of Polygonati Rhizoma and its adulterants. The developed HPLC method can be used for the differential analysis of polysaccharides in Polygonati Rhizoma and is straightforward, precise, and repeatable.

Polygonati Rhizoma  /  polysaccharides  /  enzymatic hydrolysis  /  fingerprint  /  differences  /  chemometrics  /  HPLC
刁卓, 胡冲, 杨青山, 张亚中. 基于酶解法和化学计量学的黄精多糖差异性分析*. 药物分析杂志, 2024 , 44 (2) : 324 -332 . DOI: 10.16155/j.0254-1793.2024.02.16
Zhuo DIAO, Chong HU, Qing-shan YANG, Ya-zhong ZHANG. Polygonati Rhizoma polysaccharide differential analysis based on enzymolysis and chemometrics*[J]. Chinese Journal of Pharmaceutical Analysis, 2024 , 44 (2) : 324 -332 . DOI: 10.16155/j.0254-1793.2024.02.16
黄精为2020年版《中华人民共和国药典》(简称《中国药典》)[1]收载品种,其法定基原为百合科植物滇黄精Polygonatum kingianum Coll. et Hemsl.、黄精Polygonatum sibiricum Red.或多花黄精Polygonatum cyrtonema Hua的干燥根茎,按形状不同,习称“大黄精”“鸡头黄精”“姜形黄精”,具有补气养阴、健脾、润肺、益肾等功效[2],常用于抗疲劳、调节免疫等,具有极高的药用保健价值[3-5]。由于黄精功效确切,市场认可度好,但价格较高,因此市场上常有用黄精属的其他植物,如湖北黄精、四川黄精等,冒充黄精进行销售的现象,严重影响黄精产业发展。因为黄精多糖的药材标准不够明确,采用的苯酚-硫酸法准确性低且无法获取多糖的结构信息,难以对市场上不同品种的黄精多糖质量进行有效评估,故导致这种冒充现象难以杜绝。因此,急需围绕黄精中药材多糖质量控制进行研究,以形成更加明确、更有区分度的标准。
目前对于黄精多糖的研究已经较为深入,但主要集中于采用酸水解以及柱前衍化等方法对于黄精基原物种单糖组成等方面的研究[6-8],对黄精及其混伪品的寡糖研究较少且对于黄精品种的研究较为单一。本研究基于酶解法,对于不同品种黄精特异性寡糖片段进行分析,建立了HPLC指纹图谱,并结合相似度评价以及化学计量学分析,成功实现对于黄精及其混伪品的鉴别工作,为黄精多糖的质量评价工作提供参考。
U3000型高效液相色谱仪(配备AllechELSD6000蒸发光散射检测器),Dionex公司;XP26百万分之一分析天平,Mettler Toledo公司;Elmasonic S型超声仪(功率1 kW,频率37 kHz),德祥科技有限公司;FD240电热恒温干燥箱,Binder公司;XMTD205水浴锅,常州国宇仪器制造有限公司;JW-3021 HR高速冷冻离心机,安徽省嘉文仪器装备有限公司;LGJ-10S真空冷冻干燥机,北京松源华兴科技发展有限公司;Millipore Simplicity-185型超纯水仪,Millipore公司。
对照品果糖(批号111504-201703,纯度99.8%)、蔗糖(批号111507-202105,纯度99.8%)、葡萄糖(批号110833-201908,纯度99.8%),中国食品药品检定研究院;α-淀粉酶(EC:3.2.1.1,批号S11GS160659)、果糖苷酶(EC:3.2.1.26,批号J21M11K10978),上海源叶生物科技有限公司。三氯甲烷(批号10006818)、正丁醇(批号20211026)、乙醇(批号20211026)均为分析纯,国药集团化学试剂有限公司;乙腈(批号F22M7D201)色谱级,Fisher Scientific公司;黄精药材由安徽省食品药品检验研究院提供,经安徽省食品药品检验研究院张亚中教授鉴定,分别为百合科植物滇黄精Polygonatum kingianum Coll. et Hemsl.、黄精Polygonatum sibiricum Red.或多花黄精Polygonatum cyrtonema Hua的干燥根茎。此外,文中所提到的四川黄精、湖北黄精、小叶黄精以及进口黄精为调研过程中所收集当地的习用品。34批样品详情见表1
将黄精的新鲜根茎于烘箱中烘干至恒重,并粉碎。取粉末5 g,置于15倍85%乙醇中浸泡24 h。过滤并离心(3 000 r min-1,5 min),收集沉淀并烘干,加入15倍的蒸馏水,在80 ℃条件下超声提取2次,每次2 h,离心,上清液合并,浓缩成浓度为400 mg·mL-1黄精多糖溶液。在浓缩液中加入浓度为20 mg·mL-1的α-淀粉酶,在60 ℃水浴下酶解4 h,反应完成后,沸水浴灭酶10 min。取上清液,加相当于其体积1/4的Sevage试剂,剧烈震摇,离心,重复以上操作直到无白色絮状物产生为止。所得上清液加入无水乙醇(慢加快搅)至乙醇浓度为80%,置4 ℃冰箱中放置24 h,离心,倒去上清液。沉淀加95%乙醇洗涤2次,每次10 mL,离心,得沉淀物,加热水使沉淀溶解,冷冻干燥,得黄精粗多糖冻干粉。
取黄精多糖10 mg,加水20 mL,制成质量浓度为0.5 mg·mL-1的黄精多糖溶液;另取果糖苷酶10 mg(100 U·mg-1),转移至10 mL量瓶中,用水溶解并定容至刻度,制成100 U·mL-1的果糖苷酶溶液。取黄精多糖溶液5 mL与果糖苷酶溶液(100 U·mL-1)5 mL混合。混合物在振荡器中水解(55 ℃,200 r·min-1,3 h),并在80 ℃下加热20 min使酶变性。离心(4 500 r·min-1,15 min)后,取上清液,干燥,溶解在2 mL乙腈-水(1∶1)溶液中,以进行HPLC分析。
取果糖、蔗糖、葡萄糖的对照品各约10 mg,精密称定,分别转移至10 mL量瓶中,用水溶解并定容至刻度,即得。
U3000型高效液相色谱仪(配备AllechELSD6000蒸发光散射检测器),色谱柱为Hypersil GOLDTM PEI HILIC HPLC(250 mm×4.6 mm,5 μm),流动相为乙腈(A)-水(B),梯度洗脱(0~25 min,85%A→65%A;25~35 min,65%A→50%A),流速1.0 mL·min-1,柱温35 ℃,进样量10 μL。
取S2号样品一份,按“2.2”项下方法制备供试品溶液,连续进样6次测定并记录各共有峰的相对保留时间和相对峰面积(以果糖的出峰时间和峰面积为参考,其他各峰出峰时间和峰面积与其的比值,下同),测得其共有峰相对保留时间和相对峰面积的RSD均<3.0%,表明仪器精密度良好,符合指纹图谱要求。
取S2号样品6份,按“2.2”项下方法制备供试品溶液,按“2.4”项下色谱条件进行测定,测得各共有峰的相对保留时间和相对峰面积的RSD均<3.0%,同一样品每次提取之间的结果比较一致,表明该提取方法重现性良好,符合指纹图谱要求。
取同一份S2号样品溶液,分别在0、2、4、8、12、24 h进样,测得其共有峰相对保留时间和相对峰面积的RSD均<3.0%,表明供试品溶液在24 h内较稳定,化学成分及含量不会发生改变,符合指纹图谱要求。
对上述34批样品进行HPLC分析研究,找出2020年版《中国药典》中3种不同基原黄精存在的差异及其与市场中易混品的异同,以此进行鉴别。
图1所示,HPLC检测到17个峰,得到了较为理想的特异性寡糖片段。比较相同条件下果糖、葡萄糖与蔗糖的保留时间,发现峰1、峰2、峰4分别为果糖、葡萄糖和蔗糖。此外,将5批黄精、5批滇黄精、10批多花黄精、5批四川黄精、5批湖北黄精、2批小叶黄精、2批进口黄精药材色谱图,分别导入中药指纹图谱相似度评价系统(2012版)软件,生成3种基原黄精的对照图谱。34批样品分别与黄精、滇黄精和多花黄精的对照图谱比较,计算相似度,以多花黄精对照图谱为例,如图23所示,结果见表2
从表中可看出:与多花黄精对照图谱相比,3种基原黄精的相似度中10批多花黄精相似度>0.93(图2);滇黄精和黄精与其相比(图3-A),除1批滇黄精(D1)样品外,相似度<0.85;而市场上的易混品相较于多花黄精对照图谱而言(图3-B),分布较为离散,其中湖北黄精相似度在0.44~0.95;四川黄精除1批(C4)之外,相似度均<0.86;进口黄精相似度均<0.82;小叶黄精相似度均<0.5。
上述结果表明,药典收录的3种不同来源黄精药材的种内指纹色谱相似度较高,差异较小;3种基原黄精种间差异较大。因此,不同黄精多糖的结构特征不同,便于区分。
由于相似度分析中存在部分基原黄精品种区分度不明显的情况,故进一步采用聚类分析(HCA)对黄精、滇黄精和多花黄精进行分类。取20批黄精基原物种(含滇黄精、黄精、多花黄精)中多糖的HPLC-ELSD指纹图谱的共同峰面积整合形成20×17的数据矩阵。将数据矩阵进行低级融合并导入IBM SPSS Statistic 23版软件,采用组间联接作为聚类方法,根据个体差异对样本进行分类。
HCA结果如图4所示,20批黄精样品被分为3大类:第1类包括10批多花黄精,产地分别是安徽六安、安徽池州(3个),浙江杭州(4个)、浙江衢州、江西瑞昌;第2类包括5批滇黄精,产地分别是四川达州、四川成都(2个)、湖北武汉、云南昆明和贵州毕节;第3类包括5批黄精,产地分别是贵州毕节、安徽池州、山东泰安、辽宁抚顺。结果表明,黄精多糖酶水解产物经数据分析后依次被划分为3类与基原有一定的相关性,HCA的样本中同一类别表明它们在结构特性上确实具有高度相似性。分成3类的原因可能是不同来源的黄精多糖结构具有一定的差异,这一发现给黄精的鉴别工作提供了另一种方法。
体现了降维思想的主成分分析(PCA),选取贡献率较大的成分进行分析,既简化了分析过程,又尽可能多地展现了原始信息[9]。将20批次样品的17个共有峰峰面积导入SPSS 23.软件,进行PCA,特征值和方差贡献率如表3所示,碎石图见图5。以特征值>1为提取标准[10],提取了4个主成分,前3个分别占总方差的50.1%、26.5%和9.1%,故选取3个主成分进行评价,它代表黄精多糖中17个成分量的85.8%的信息量,足以评价黄精多糖的品质。将数据矩阵导入SIMCA统计软件中,提取了3个主成分,该模型中,R2X(cum)=0.844,Q2(cum)=0.612,均>0.5,说明该PCA模型的预测能力越好。黄精多糖的三维PCA得分图(图6)表明,20批黄精多糖被分为3类,与HCA结果一致。多花黄精S1~S10被聚集为一组,滇黄精D1~D5被聚集到另一组,而鸡头黄精J1~J5被分到第3组。PCA模型也为3种不同基原的黄精提供了令人满意的分类,表明采取酶水解的方法对于黄精多糖进行分析,可以对不同来源的黄精进行很好的鉴别。
将20×17的数据矩阵导入SIMCA 13统计软件进行偏最小二乘判别分析(PLS-DA),每个色谱峰对鉴别结果的影响由VIP(投影中的变量重要性)值确定。该模型中,R2X(cum)=0.909,R2Y(cum)=0.863,Q2(cum)=0.775,R2YQ2的值以>0.5为宜,且R2YQ2的值越趋近于1,说明该PLS-DA模型的预测能力越好[11]。分析结果如图7所示,分类结果与HCA和PCA相一致。图8反映了该模型的VIP值,VIP值的大小表示的是每个自变量贡献的大小,以VIP>1为筛选标准,得出导致品种间差异的标志性成分。从VIP值可知,色谱峰1、4、5、7和8的VIP值都>1,说明上述5个色谱峰对PLS-DA模型的判别分析具有较大的贡献。
根据文献调研发现,部分酸水解已广泛应用于中药多糖的降解[12-13],但基于酶解的方式对中药多糖进行研究更具方向性、目的性和可控性[14],在分析多糖的糖苷键类型、构建糖谱以及构效关系研究等方面具有显著优势。虽也存在反应条件要求较高,反应时间长,合适的酶不易获取等问题,但是现已完成适用于黄精多糖的特异性糖苷酶筛选以及酶解条件的优化,且还没有基于酶解法对于不同品种黄精多糖研究的报道。黄精基原混乱的主要原因是目前市售黄精多出自于人工栽培,性状往往与基原物种极其相似。
本实验采用果糖苷酶对于黄精多糖展开研究,通过相似度评价鉴别不同品种黄精之间的差异,结果表明果糖苷酶水解下3种基原黄精的种内指纹色谱相似度较高,差异较小,而种间差异较大。特别是以多花黄精为对照图谱时结果较为明显,滇黄精和黄精与其相比,除1批滇黄精(D1)样品外,相似度均<0.85。而市场上的易混品相较于多花黄精对照图谱而言,分布较为离散,其中湖北黄精相似度在0.44~0.95,这可能是由于市场上存在多花黄精掺伪湖北黄精的情况。四川黄精除1批(C4)外,相似度均<0.86,提示四川黄精与多花黄精依旧存在差异,故而市场上将其与多花黄精混为一谈的说法有待进一步验证。此外,进口黄精相似度均<0.82,小叶黄精相似度均<0.5,表明不同黄精多糖的结构特征不同。此方法一定程度上解决了目前市场上对于四川黄精、湖北黄精等黄精易混品鉴别困难等问题。
本实验在仪器的使用方面,选择了灵敏的蒸发光检测器,更适用于对于多糖中寡糖片段的分离[15]。通过近一步与HILIC色谱柱相结合,最终黄精多糖指纹图谱可达到17个色谱峰,对于大分子糖的研究有了新的突破,通过定位酶切技术成功得到特异性寡糖片段。后续对于药典收录的黄精品种,采用化学计量学分析,通过PCA发现,采用果糖苷酶可以很好地按照黄精种类的不同将其进行分类,而VIP值结果表明,峰1、4、5、7和8可以作为区分不同品种黄精的标志物,这也再次验证了采用酶解方法结合HPLC分析黄精多糖的可行性,进一步推动黄精多糖的研究应用与快速发展。
  • *安徽省药品监督管理局监管科学研究重点项目-常用中药材DNA分子鉴定研究
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2024年第44卷第2期
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doi: 10.16155/j.0254-1793.2024.02.16
  • 接收时间:2023-06-19
  • 首发时间:2026-03-13
  • 出版时间:2024-02-29
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  • 收稿日期:2023-06-19
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*安徽省药品监督管理局监管科学研究重点项目-常用中药材DNA分子鉴定研究
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    1.安徽医学高等专科学校,合肥 230031
    2.安徽省食品药品检验研究院 国家药监局中药质量研究与评价重点实验室,合肥 230051
    3.安徽中医药大学,合肥 230012

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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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