丹参及丹参茎叶水提物UPLC指纹图谱研究与丹酚酸类成分定量分析

目的：建立并比较丹参、丹参茎叶水提物UPLC特征指纹图谱，同时分析评价7种丹酚酸类成分（丹参素、原儿茶醛、咖啡酸、迷迭香酸、紫草酸、丹酚酸B和丹酚酸A），以期为丹参资源的综合利用提供科学依据。方法：采用ACQUITY UPLC BEH C₁₈色谱柱（2.1 mm×100 mm，1.7 μm）；流动相为乙腈（A）-0.1%甲酸水溶液（B），梯度洗脱（0~1 min，5% A；1~3 min；5% A → 10% A；3~7 min，10% A → 15% A；7~11 min，15% A → 21% A；11~15 min，21% A → 33% A；15~17 min，33% A → 70% A；17~18 min，70% A → 80% A；18~20 min，80% A；20~21 min，80% A → 5% A），流速0.4 mL·min^-1；检测波长为280 nm。采用“中药色谱指纹图谱相似度评价系统”建立对照图谱和进行相似度评价，SPSS 19.0进行主成分分析和聚类分析。结果：丹参素、原儿茶醛、咖啡酸、迷迭香酸、紫草酸、丹酚酸B和丹酚酸A线性范围分别为1.145~229、0.81~162、0.95~190、1.265~253、1.285~257、1.055~211和0.87~174 μg·mL^-1，R²为0.999 8以上；平均回收率在97.3%~103.5%之间，RSD均小于3%（n=6）。结论：主成分分析将丹参、丹参茎叶各聚为一类，聚类分析和相似度评价表明丹参及其茎叶水提物指纹图谱具有一定相似性，相似度最高可达0.981，共有化学成分主要为丹酚酸类成分。定量分析表明不同批次丹参、丹参茎叶中丹酚酸类成分含量存在一定差异，但丹酚酸类成分的种类组成相一致。

Objective: To establish an ultra-high performance liquid chromatography method to compare the characteristic fingerprint of Radix et Rhizoma Salviae Miltiorrhizae and stems and leaves of Salvia miltiorrhiza. And to determine the contents of seven kinds of salvianolic acids(danshensu, protocatechuic aldehyde, caffeic acid, rosmarinic acid, lithospermic acid, salvianolic acid B and salvianolic acid A). To provide scientific basis for the comprehensive utilization of Salvia miltiorrhiza resource. Methods: The separation was performed on an ACQUITY UPLC BEH C₁₈(2.1 mm&#215;100 mm, 1.7 μm) column with gradient elution of acetonitrile(A) and 0.1% formic acid(B) (0-1 min, 5%A; 1-3 min; 5%A → 10%A; 3-7 min, 10%A → 15%A; 7-11 min, 15%A → 21%A;11-15 min, 21%A → 33%A;15-17 min, 33%A → 70%A;17-18 min, 70%A → 80%A;18-20 min,80%A;20-21 min,80%A → 5%A). The flow rate was 0.4 mL&#183;min^-1 and the UV detection were at 280 nm. Similarity evaluation system for chromatographic fingerprint of TCM was used for establishing the reference fingerprint and similarity evaluation, and the principal component analysis(PCA) and clustering analysis was carried out by SPSS 19.0. Results: The calibration curves of danshensu,protocatechuic aldehyde, caffeic acid, rosmarinic acid, lithospermic acid, salvianolic acid B and salvianolic acid A were all in good linearity over the ranges of 1.145-229, 0.81-162, 0.95-190, 1.265-253, 1.285-257, 1.055-211 and 0.87-174 μg&#183;mL^-1, respectively,with correlation coefficients R² over 0.999 8. The average recoveries ranged from 97. 3% to 103.5%, with RSD(n=6)<3.0%. Conclusion: It showed that PCA clustered Radix et Rhizoma Salviae Miltiorrhizae and stems and leaves of Salvia miltiorrhiza for one class by themselves. Cluster analysis and similarity evaluation indicated that in some way,fingerprint of water extracts of Radix et Rhizoma Salviae Miltiorrhizae and stems and leaves of Salvia miltiorrhiza were similar, with maximum similarity up to 0.981. Salvianolic acids were identified to be the common components. Results of assay showed that salvianolic acids in different batches of Radix et Rhizoma Salviae Miltiorrhizae and Salvia stems and leaves varied a lot in contents but kept consistent in compositions.

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