不同产地山楂叶HPLC指纹图谱比较分析

目的 建立不同产地的山楂叶50％乙醇提取物的HPLC指纹图谱,并对其中5种主要成分进行含量测定,结合聚类分析评价不同产地的山楂叶药材质量.方法 采用Diamonsil C18(4.6 mm×250 mm,5μm)色谱柱;以乙腈-0.1％冰醋酸-四氢呋喃体系梯度洗脱;检测波长320 nm;流速0.9 mL·min-1;柱温30℃;应用中药色谱指纹图谱相似度评价系统建立HPLC指纹图谱进行评价,用SPSS 19.0统计软件进行聚类分析.结果 标定出山楂叶HPLC指纹图谱共有峰7个,相似度0.09～0.90;聚类分析后,可以区分不同产地山楂叶,最终以19批主产地的山楂叶为样本,重新建立山楂叶50％乙醇提取物HPLC指纹图谱,标定出共有峰11个,相似度均大于0.85;对25批山楂叶提取物5种主要成分进行含量测定,结果显示主产区山楂叶的质量比较稳定.结论 建立的山楂叶HPLC指纹图谱重现性和特征性较好,能够快速鉴别山楂叶.应用SPSS 19.0聚类分析软件,可以较全面的反映不同产地山楂叶化学成分的差异,为其质量控制提供实验依据.     

大蓟不同药用部位的HPLC特征图谱与化学模式识别研究

目的:建立大蓟不同药用部位的高效液相色谱(HPLC)特征图谱,并结合化学识别方法全面比较大蓟不同药用部位化学成分的差异,为大蓟药材的质量控制及评价提供参考。方法:采用HPLC法分别对大蓟药材(即地上部分)、叶、花、主茎和侧茎进行测定,按《中药色谱指纹图谱相似度评价系统》(2012A版)对色谱图进行匹配生成各药用部位的HPLC特征图谱,通过单因素方差分析共有特征峰峰面积差异,通过与对照品比对进行色谱峰指认,并结合主成分分析和聚类分析对大蓟不同药用部位进行化学模式识别研究。结果:分别建立了大蓟药材、叶、花、主茎、侧茎的HPLC特征图谱,大蓟药材、叶和花的图谱中共确定了15个共有峰,主茎、侧茎的图谱中共确定了11个共有峰(7、9、12、13号峰缺失),不同药用部位之间化学成分含量差异较大;指认峰1、2、3、10、11分别为新绿原酸、绿原酸、隐绿原酸、蒙花苷和柳穿鱼叶苷。主成分分析和聚类分析结果显示,大蓟花与茎的化学模式识别聚类区别清晰,可分别聚为一类;但大蓟叶分布较为分散,难以很好地聚类。结论:建立的HPLC特征图谱-化学模式识别分析模型能够整体、全面、真实地反映大蓟不同药用部位之间的差异,...     

不同产地和品种天麻的HPLC指纹图谱研究

目的 研究不同产地和不同品种天麻的HPLC指纹图谱.方法 色谱柱为Diamonil C18柱(250mm×4.6 mm,5μm),流动相为甲醇-0.1％磷酸水溶液,梯度洗脱,检测波长220 nm,流速1 mL·min-1,柱温35℃.对20批天麻样品的HPLC指纹图谱进行相似度评价和聚类分析.结果 确立了13个共有峰,建立了天麻药材HPLC指纹图谱共有峰模式.20批不同产地和品种天麻的相似度为0.829 ～1.000,综合评判得分为-0.77～1.07.结论 建立的HPLC指纹图谱反映了天麻药材中的化学成分信息,同一产地天麻的化学成分趋同,而不同产地的天麻差异较大,表明产地因素对其化学成分的影响较大.     

柴胡高效液相色谱数字化指纹图谱研究

目的用反相高效液相色谱法建立柴胡HPLC数字化指纹图谱。方法采用CenturySIL C18 AQ色谱柱(20 cm×4.6 mm,5μm),流动相为水和乙腈(均含1%醋酸)梯度洗脱;流速1.0 mL.min-1;265 nm检测;柱温(30±0.15)℃。采用“中药色谱指纹图谱超信息特征数字化评价系统”软件进行评价。结果以尿苷峰为参照物峰,确定了27个共有峰。通过对指纹峰分离程度、信号响应强度、信号均化性、信息量多寡等多种参数评价,建立了柴胡HPLC数字化指纹图谱。各产地药材的化学成分分布比例和含量2个方面与对照指纹图谱间都具有良好相似性。结论本法可清晰揭示柴胡HPLC指纹图谱的超信息特征,方法具有较好的稳定性和重现性,为柴胡质控提供了新参考。     

三七高效液相色谱数字化指纹图谱研究

目的建立三七HPLC数字化指纹图谱。方法采用RP-HPLC法,CenturySIL C18BDS(250 mm×4.6mm,5μm),流动相为水-乙腈梯度洗脱,流速0.9 mL.min-1。紫外检测波长203 nm,柱温(40.0±0.15)℃,进样量5μL。以"中药色谱指纹图谱超信息特征数字化评价系统"软件进行评价。结果以人参皂苷Rb1峰为参照物峰,确定20个共有峰,建立了三七HPLC数字化指纹图谱。应用色谱指纹图谱指数F等参数对不同批次三七的HPLC指纹图谱的超信息特征进行了数字化评价。结论所建立的HPLC数字化指纹图谱具有较好的精密度、重现性和稳定性,适用于三七药材的质量控制。     

中药皂角刺的HPLC指纹图谱

目的建立不同产地皂角刺HPLC指纹图谱,为皂角刺质量评价提供科学依据。方法采用RP-HPLC法,色谱柱为Agilent ZORBAX SB-Aq C_(18)(250 mm×4.6 mm,5μm),流动相为甲醇(A)-体积分数为0.1%的乙酸溶液(B),梯度洗脱,检测波长为254 nm,柱温为35℃,流速为1.0 m L·min-1。采用中药色谱指纹图谱相似度评价系统(2012.130723版本)和SPSS 21.0软件对谱图分别进行相似度分析和聚类分析。结果建立了皂角刺HPLC指纹图谱,确定了29个共有峰,并指认了其中5个共有峰的化学成分。从HPLC指纹图谱结果看,不同产地皂角刺化学成分相似,但含量差异较大。结论中药皂角刺HPLC指纹图谱可为中药皂角刺质量评价提供科学依据。     

贵州产杜仲高效液相色谱指纹图谱的建立与分析

目的 建立杜仲药材高效液相色谱(HPLC)的指纹图谱,评价不同产地杜仲中化学成分的差异,为制定杜仲的质量控制标准提供参考.方法 色谱柱为Phenomenex Synergi Hydro-RP C18(250 mm×4.6 mm,4μm),流动相为甲醇-0.05％磷酸溶液(梯度洗脱),检测波长235 nm,柱温25℃.利用中药色谱指纹图谱相似度评价系统软件进行相似度计算,并应用聚类分析与主成分分析对指纹图谱进行化学模式识别法研究.结果 建立了杜仲药材的指纹图谱,确定了19个共有峰,并用对照品指认了3个峰.相似度评价表明,20批不同产地的杜仲药材相似度存在差异;聚类分析将20个样品分成A与B两类.结论 杜仲药材指纹图谱的建立为杜仲的质量控制评价体系提供了科学依据.     

广西产赪桐的HPLC指纹图谱研究

目的 研究广西产赪桐的HPLC指纹图谱.方法 采用Hypersil C18色谱柱(250 mm ×4.6 mm,5μm),流动相为0.1％磷酸-甲醇,梯度洗脱,检测波长320 nm,柱温25℃,流速1.0 mL· min-1;将色谱图导入《中药色谱指纹图谱相似度评价系统(2004A版)》软件中,生成对照图谱,并进行相似度评价和聚类分析.结果 建立了含12个共有峰的赪桐特征指纹图谱,不同产地赪桐特征图谱的相似度均＞0.9.结论 所建指纹图谱反映了同一产地赪桐中化学成分的趋同,可用于赪桐药材的质量控制.     

龙眼叶乙酸乙酯部位的HPLC指纹图谱研究

目的建立不同产地龙眼叶药材乙酸乙酯部位的HPLC指纹图谱,并进行聚类分析和主成分分析。方法采用Thermo C18(4. 6 mm×150 mm,5μm)色谱柱,甲醇-0. 2%磷酸水溶液为流动相梯度洗脱,流速1. 0 mL·min-1,柱温30℃,检测波长280 nm。结果提取12个色谱峰为药材指纹图谱共有峰,采用对照品指认了3个主要色谱峰,其中以对照品没食子酸乙酯为参照峰,运用相似度评价软件、聚类分析和主成分分析对所收集的10个不同产地龙眼叶药材乙酸乙酯部位进行系统比较与归类,10个产地龙眼叶药材相似度均大于0. 9,分别聚为4类。结论建立的指纹图谱方法重复性好,简便可靠,为不同产地地龙眼叶药材质量控制和评价提供依据。     

印楝叶HPLC指纹图谱及其化学成分与遗传变异的关系研究

目的:建立印楝叶的高效液相色谱(HPLC)指纹图谱,并分析其化学成分与遗传变异的关系。方法:采用HPLC法进行指纹图谱研究,运用简单序列重复间扩增多态性(ISSR)技术分析遗传多样性,利用聚类分析研究二者的关系。结果:建立了印楝叶药材的HPLC指纹图谱,共标定了11个共有指纹峰。分别根据Nei’s遗传距离和化学成分进行聚类分析,13个种源分为两大类:来自缅甸的11个种源聚成一类;来自澳大利亚和印度的种源聚成另外一类。结论:所建立的指纹图谱稳定、可靠、重复性好;不同种源印楝叶药材化学成分与遗传变异之间具有一定的相关性。     

Detection of genetic homogeneity of Panax notoginseng cultivars by sequencing nuclear 18S rRNA and plastid matK genes

The nuclear 18S rRNA and chloroplast MATK genes of 18 samples of Panax notoginseng and its processed material Sanqi (Radix Notoginseng) were analyzed. The two genes, regardless of cultivar origin, were found to be identical to genotype R1 and M1, respectively, of the published sequences (GenBank accession no. D85171 and AB027526). This phenomenon implies that the species is highly conserved, which is probably caused by the use of the same strain in cultivation and the lack of active mutation in these two genes.     

Genetic heterogeneity of ribosomal RNA gene and matK gene in Panax notoginseng

Previously, 185 ribosomal RNA gene and matK gene sequences of Chinese herbal medicines, Ginseng Radix, Panacis Japonici Rhizoma and Panacis Quinquefolli Radix were shown to correspond with those of the original plants, Panax ginseng, P. japonicus and P. quinquefolius, respectively, with the species-specific sequences especially for 18S rRNA gene sequences. In P. notoginseng and its derivative, Notoginseng Radix, however, we found two genetic groups with respect to both gene sequences. Five base substitutions were detected on both gene sequences and the homology between two groups was 99.7% for the 18S rRNA gene and 99.6% for the matK gene, respectively. One genetic group was found to have the identical sequences as those of P. ginseng.     

Authentication of Panax notoginseng by 5S-rRNA spacer domain and random amplified polymorphic DNA (RAPD) analysis

The great majority of Panax species are well-known herbal medicines in the Orient, and many of them share a close resemblance in appearance and chemical composition. Among these Panax species, the root of P. notoginseng (Sanqi) is a unique herb that has distinct clinical usage. Here, the 5S-rRNA spacer domains were isolated from P. notoginseng, P. japonicus var. major, P. stipuleanatus, P. quinquefolius, P. ginseng, P. zingiberensis, and P. wangianus, and four common adulterants of P. notoginseng including Curcuma wenyujin, Curcuma longa, Bletilla striata and Gynura segetum. The spacer domains were sequenced and compared, which showed over 75 % DNA identity among all Panax species, but not for the adulterants. In addition, random amplification of polymorphic DNA (RAPD) analysis was used to distinguish different members of Panax genus as well as the morphological variants of P. notoginseng. These molecular methods could be used in the authentic identification of P. notoginseng from other Panax species.     

Species identification from Ginseng drugs by multiplex amplification refractory mutation system (MARMS)

The multiplex amplification refractory mutation system (MARMS) was applied to the identification of 5 Panax species ( P. ginseng, P. japonicus, P. quinquefolius, P. notoginseng and P. vietnamensis). A set of specific primers, including 2-pair primers on chloroplast trnK gene and nuclear 18S rRNA gene regions, respectively, was designed and synthesized for each species on the basis of species-specific sequences of the 2 genes. By using 5 sets of specific primers, in turn, PCR amplifications were performed with total DNA extracted from 5 Panax species as template under appropriate condition, and each resulting product was detected by agarose gel electrophoresis. The results showed that two expected fragments, one from trnK gene and another from 18S rRNA gene regions, were observed simultaneously only when the set of species-specific primers encountered template DNA of the corresponding species. This assay could give more reliable results for identification of not only 5 Panax species but also corresponding Ginseng drugs by simultaneous detection of 4-site nucleotide differences on 2 completely different genes.     

Binary chromatographic fingerprint analysis of Stemonae Radix from three Stemona plants and its applications

The dried root tubers of Stemona tuberosa, S. japonica and S. sessilifolia are the original sources of Stemonae Radix (SR) for antitussive and insecticidal activities. The products of SR which are available on the market are variable, and imitations exist. In order to characterize the overall chemical constituents of SR and evaluate its quality, a novel, binary high-performance liquid chromatographic fingerprinting method, describing the pattern of alkaloids (fingerprint I) and non-alkaloids (fingerprint II) of SR was developed. It was also applied to determine whether the medicinal parts and the processing methods affect the quality of SR. Similarity and high-performance liquid chromatography?mass spectrometry (HPLC–MSⁿ) were utilized to compare or identify the chemical constituents of SR. The results indicate that the chemical constituents from different parts of the underground material of Stemona plants are diverse and that the processing methods affect certain constituents in the root tuber samples. The similarity and the resulting chemical consitituents obtained show that the binary chromatographic fingerprint method can be used to differentiate the three official Stemona species or the adulterants of SR, which is helpful for the identification and quality evaluation of SR.

     

中药三七高效液相色谱特征研究

目的建立中药三七的高效液相色谱特征谱。方法运用加压溶剂提取、HPLC-DAD分析28个不同产地的三七药材,所得的图谱采用中国药典委员会开发的“中药色谱指纹图谱相似度评价系统（Version 2004A）”软件进行分析。结果28个三七样品色谱图中各色谱峰分离良好,从中确定了13个特征峰,其中8个主要色谱峰经对照品确认,所有药材色谱图与软件生成的对照谱相似度为0.982±0.008(RSD=0.78%)。结论该方法简便、重现性好、具可操作性,可科学评价及有效控制三七药材质量。     

Phytochemical and analytical studies of Panax notoginseng (Burk.) F.H. Chen

Panax notoginseng (Burk.) F.H. Chen is distributed throughout the southwest of China, Burma and Nepal. The root of this plant, called notoginseng or sanchi, has a long history of use as a remedy in Oriental traditional medicine. Modern studies have found that extracts and compounds from notoginseng exert various physiological effects. The active constituents are mainly recognized as saponins. In this review, we summarized the discovery and analysis of chemical constituents in notoginseng. Fifty-six saponins from notoginseng were isolated and elucidated. All of them are dammarane saponins, 35 of which can be classified as belonging to the protopanaxadiols group, and 21 as belonging to the protopanaxatriols group. Evidence from phytochemical studies on notoginseng demonstrated that no oleanane-type saponin, which exists in Asian ginseng (Panax ginseng) and American ginseng (Panax quinquefolius), was found. Other types of compounds such as non-protein amino acids, polyacetylenes, phytosterols, flavonoids, and polysaccharides, many of which have pharmacological activities, were also isolated from notoginseng. Analytical studies on notoginseng were carried out based on botanical and phytochemical advances. In the qualitative studies, identification of the herbal materials and extracts was the main objective. The utilization of high-performance liquid chromatography (HPLC) fingerprint and molecular biological methods made the identification accurate and efficient. Spectral, chromatographic and immunoassay methods were used for the quantitative analysis. HPLC methods are the main authority regarding the determination of saponins and other types of constituents. The chromatographic conditions and detectors employed in the HPLC are discussed.     

中药三七对大鼠心肌缺血保护作用的谱效学研究

目的 以大鼠心肌缺血模型为对象,研究中药三七对大鼠心肌缺血保护作用谱效关系及药效物质基础。方法 在建立中药三七液相指纹图谱分析方法的基础上,研究药物治疗心肌缺血作用的物质基础及谱效关系。结果 人参皂苷Rg₁、人参皂苷Rb₁、三七皂苷R₁是中药三七治疗心肌缺血的主要有效成分。结论 通过谱效关系研究直接获得药效活性物质,建立了三七药材谱效关系评价的新方法,客观反映了药物的内在质量,为该类中药的进一步研究提供了新的思路。     

三七主要皂苷类药效成分含量的近红外光谱测定法

目的 建立基于近红外光谱的快速测定三七药材中五种主要皂苷类药效成分含量的方法。方法 取173批三七不同部位、不同产地、不同大小规格的药材,采用HPLC定量分析方法测定三七皂苷R1、人参皂苷Rg1、Re、Rb1和Rd的含量,作为参考值。药材粉碎后在4000~10000 cm-1波数范围内采集光谱,对光谱预处理方法、建模波段及主成分数进行优选,采用偏最小二乘回归算法建立近红外光谱与三七药材中五种主要药效成分含量HPLC分析结果之间的多元校正模型。结果 三七皂苷R1、人参皂苷Rg1、Re、Rb1和Rd在校正模型中的预测相关系数（r）分别为0.9596、0.9785、0.9026、0.9660和0.9929。结论 该方法测定的五种皂苷是三七的主要药效成分,且为三七总皂苷类制剂的质量检测指标。本方法操作简便快速,结果准确,可用于三七药材质量的快速检测。     

Simultaneous determination of nine saponins from Panax notoginseng using HPLC and pressurized liquid extraction

A HPLC and pressurized liquid extraction (PLE) method was developed for simultaneous determination of nine saponins, including notoginsenoside R1, ginsenoside Rg1, Re, Rf, Rb1, Rc, Rb2, Rb3 and Rd in Panax notoginseng. The analysis was performed on C18 column with water-acetonitrile gradient elution and the investigated saponins were authenticated by comparing retention time and mass spectra with their reference compounds. Several methods including PLE, ultrasonication, soxhlet extraction and immersion were used for sample preparation and their extraction efficiency was compared. The results showed that PLE has the highest extraction efficiency and repeatability, which would be valuable on standardization of sample preparation for quality control of Chinese medicines. The developed HPLC and PLE is an effective approach for simultaneously quantitative determination of sapoinins in P. notoginseng, which could be used for quality control of P. notoginseng and its preparations.